Presentation on theme: "SFA 2073 Topic II Amino Acid & Proteins Nik Norma Nik Mahmood (PhD) Faculty Science & Technology Uni.Science Islam Malaysia NILAI, N.Sembilan."— Presentation transcript:
SFA 2073 Topic II Amino Acid & Proteins Nik Norma Nik Mahmood (PhD) Faculty Science & Technology Uni.Science Islam Malaysia NILAI, N.Sembilan
OBJECTIVES To Classify amino acids according to their structures and properties. To explain the meaning of pKa and pI of amino acids To understand the biochemical benefit of amino acids To describe the importance of some amino acids in the synthesis of important compounds To understand the biochemical benefit of proteins To Classify proteins according to their structures and properties. Relate the structure of proteins to their functions using specific examples. To understand the importance of amino acids & protein in biochemical efficiency.
INTRODUCTION: Expression of Concentration (the various expressions of concentrations used). At the end of this lecture, students should be able to: Differentiate molarity and molality Apply the units of concentration used in medicine (g%, mmol, g/dl, IU/I etc.) Explain dilution, concentrated, saturated and supersaturated solution Explain biological solution concentration ie hypertonic, hypotonic and isotonic.
Solution I. There are several way to represent concentration of solution: a) Molarity (M) the number of moles of solute per liter solution. Unit: or or molar (M) b) Molality (m) the number of moles of solute per kg of solvent. Unit: molal (m) or molkg -1 M = moles of solute (mol) Volume of solution (dm 3 or liter) moldm -3 molL -1 m = moles of solute (mol) mass of solvent (kg)
II. Units of concentration used in biological science: a) Percent Composition by Mass (%) Ratio of the mass of solute to the mass of solution multiply by 100. eg 20g NaCl in 100 g salt solution 20 x 100 = 20 % sodium chloride solution 100 b) mmol: millimol = 1X mol or 1 mol= 10 3 millimol c) g/dl : gdl -1 = g in 1 deciliter solution 10 dl = 1 L 1 dl = L
d) IU/I : International Unit is a unit of measurement for the amount of a substance, based on measured biological activity or effect. The unit is used for vitamins, hormones, some medications, vaccines, blood products, and similar biologically active substances. IU is not part of the International System of Units used in physics and chemistry. IU should not be confused with the enzyme unit, also known as the International unit of enzyme activity and abbreviated as U.enzyme unit Mass equivalents of 1 IU Insulin: 1 IU is the biological equivalent of about 45.5 μ g pure crystalline insulin (1/22 mg exactly) Insulin μ g Vitamin A: 1 IU is the biological equivalent of 0.3 μ g retinol, or of 0.6 μ g beta-carotene Vitamin Aretinolcarotene Vitamin C: 1 IU is 50 μ g L-ascorbic acid Vitamin Cascorbic acid Vitamin D: 1 IU is the biological equivalent of μ g cholecalciferol/ergocalciferol Vitamin D cholecalciferolergocalciferol Vitamin E: 1 IU is the biological equivalent of about mg d-alpha- tocopherol (2/3 mg exactly), or of 1 mg of dl-alpha-tocopherol acetate Vitamin Emg tocopherol
III. a) Making Dilutions III. a) Making Dilutions process of adding more solvent to a known solution. The moles of solute stay the same, moles = M x L In solution: initial Mole of solute = final Mole of solute M 1 V 1 = M 2 V 2
III. b) concentrated solution has less amount of water and more amount of the substance. For example concentrated H 2 SO 4 has 2% water and 98% H 2 SO 4 and dilute has less amount of substance and more amount of water c) saturated solution contains the maximum amount of a solute that will dissolve in a given solvent at a specific temperature. d) supersaturated solution contains more solute than is present in a saturated solution at a specific temperature. e) biological solution: concentration is described as hypertonic or hypotonic Hypertonic solution contain a high concentration of solute relative to another solution on the other side of the membrane. Water from the other side will flow to this solution.
Few notes for weak acid: pH is a direct measure of the H + concentration. Ka: is acid dissociation/extent of ionisation constant, acidity constant. pKa:The negative logarithm of K a pKb:The negative logarithm of the base protonation constant K b the extent of ionization of a weak acid (the pKa) influences the final concentration of H + ions (the pH) of the solution. For a weak acid there is a relationship between pH and its pKa. This relationship is given by the Henderson–Hasselbalch equation: pKa = pH + log [HA] / [A-] OR pH= pKa + log [A-] / [HA] can be CH 3 CO 2 - CH 3 CZRCO 2 -
Derivation of Henderson–Hasselbalch equation K a = [H 3 O + ] [CH 3 COO - ] [CH 3 COOH] [H 3 O + ] = K a [CH 3 COOH] [CH 3 COO - ] X each by (- log) …… – log [H 3 O + ] = – log K a – log [CH 3 COOH] [CH 3 COO - ] pH = pK a – log [CH 3 COOH] [CH 3 COO - ] pH = pK a + log [CH 3 COO - ] [CH 3 COOH] Henderson–Hasselbalch equation.
Determination of pKa Titration of 100 mL 0.1 M CH 3 CO 2 H with 50 mL 0.1M NaOH CH 3 CO 2 H + NaOH CH 3 CO 2 + Na +H 2 O stoichiometric coefficient 1:1 Initial mole CH 3 CO 2 H = 0.01 Final mole CH 3 CO 2 ˉ =0.005 Unreacted CH 3 CO 2 H = = pH value can be determined by using pH meter Substituting all the values in the equation, can get pKa By varying the volume of 0.1M NaOH in each titration can get the corresponding pH and pKa values
Relationship between amino acids and protein: Amino acids are building units of protein Peptide bonds Different coloured balls & box => Amino Acids Protein n
Amino Acid: Structure & Function Amino acid (a.a) 20 altogether = std aa - - all aa share a general formula R-CH-NH aa differ from the other by the feature of –R - - Classified based on : i) structure ii) side chain aliphatic aa non-polar dicarboxylic aa uncharge or non- ionic polar diamino aa charge or ionic aromatic aa polar heterocyclic aa COOH
Aliphatic Non-polar Amino Acid hydrophobicity Properties: - glycine and alanine are also found in the free form.
Aromatic Amino Acid Properties: tryptophane phenylalanine -Are non polar - absorb ultraviolet light (to different degree) - tyrosine has ionizable side chain
Basic Amino Acid Properties: - Are polar - Are positively charged at pH values below their pKas -Are very hydrophilic - imidazole of histidine, at pH 7 exist predominantly in the neutral form. Histidine lysine arginine
Acidic Amino Acid Properties: -are polar - are negatively charge at physiological pH - the –COOH of side chain can form amide with an amino group.
EssentialNonessential IsoleucineAlanine LeucineAsparagine LysineAspartate MethionineCysteineCysteine* PhenylalanineGlutamate ThreonineGlutamineGlutamine* TryptophanGlycineGlycine* ValineProlineProline* SerineSerine* TyrosineTyrosine* ArginineArginine* HistidineHistidine* * Essential in certain cases. Eg arginine & histidine are growth promoting factor there fore become essential in growing children - iii) Nutritional Requirement essential aa (8/9). Cannot be synthesized by the body non-essential aa (12/11). Can be synthesized by the body
- Amino acid is a derivative of organic (weak) acid. - Has 2 functional groups, carboxylic group (-COOH) and amino group (-NH 2 ). Carboxylic (-COOH) and amine (-NH 2 ) groups are capable of ionization: COOH COO + H + (2< pKa1< 2.5) N + H 3 NH 2 + H + (9< pKa2< 9.5) ( N + H 3 is a weaker acid ) - All aa is affected by pH: The net charge on the molecule in solution is affected by pH of their surrounding and can become more positively or negatively charged due to gain or the loss of protons (H + ) respectively. eg. At pH~2.0 the amino group will be as –NH 3 +, the carboxylic group will remain as –COOH (aa will migrate towards the cathode). As pH is increased, –COOH (from some fraction of aa) ionises. When the pH is equal to the pKa1 the amino acid exists as a 50:50 mixture of the cationic and zwitter ionic forms. As pH is further increased more cationic form converts to the zwitterionic Can donate & accept H + i.e amphoteric nature therefore aa are ampholytes
- Adding more base results in continued ionization of the carboxylic acid group until the zwitter ionic form is the predominant form of the amino acid in solution. By the addition of more base, the pKa of the amino group is reached and at this point the amino acid exists as a 50:50 mixture of the zwitter ionic form and the anionic form. As the pH is increased further the amino group continues loses its proton and ultimately, at high pH (pH ~ 12.0), the anionic form is the predominant form in solution. At pH>~9.6 the amino group will be as –NH 2, the carboxylic group will remain as -COOˉ (aa will migrate towards the anode). - - So at physiological pH , the –COOH group exist as COO¯, and the –NH 2 as –NH 3 +. Therefore all aa are double-charged structure or zwitterion in this pH region. The pH at which they exist as whole zwitterion i.e the molecule carries no electrical charge, or the negative and positive charges are equal is called Isoelectric point (Ip) or Isoelectric pH.
- Each aa has its Ip value. At Ip: i) aa is double-charge (zewtterionic) i.e +ve & -ve, amount of positive charge exactly balances the amount of negative charge so net charge is 0 (electrically neutral). ii) it does not move/migrate in electric current iii) the molecule has minimum solubility. iv) Ip of all aa lie in the range of pH Isoelectric pH of an aa solution is given by: pH = ½ (pK 1 + pK 2 ) CH 3 -CHCOOH CH 3 CH COO¯ Neutral un-charged NOT THIS NH 2 N+H3N+H3 Zwitterion. Neutral but charge aa Actual structure Low pH region High pH region
The pH profile of an acidic solution of alanine when the solution is titrated with a strong base, NaOH. E.g 50% as cationic 50% as zwitterion 50% as anionic 50% as zwitterion For aliphatic aa
Physical properties: - colourless crystalline; soluble in water/polar solvents. Tyrosine is soluble in hot H 2 O - have high m.pt >200 o C - have high dielectric constant and high dipole moment - molecules have minimum solubility in water or salt solutions at the Ip pH and often precipitate out of solution.Why? At Ip aa is in zwitterionic form therefore non-polar. Hence no interaction with polar water molecules Chemical properties: involve –COOH & involve –NH 2 i) involve –COOH decarboxylation or formation of amine & CO 2 eg. histadine histamine + CO 2 tyrosine tyromine + CO 2 tryptophan tryptamine + CO 2 lysine cadaverine CO 2 Glutamic gamma amino butyric acid (GABA) + CO 2
Amide formation : α-COOH of 1 aa reacts with α-NH 2 of aa behind to form a peptide bond or CONH bridge eg in peptides and proteins Amide formation (at 2 nd COOH) aspartic + NH 3 asparagine glutamic + NH 3 glutamine (than N donated for N.A synthesis) ii) involve –NH 2 : formation of carbamino compound –NH 2 + CO 2 –NH-CO 2 H eg transport of CO 2 by hemoglobin from tissue to lung Hb–NH 2 Hb–NH-CO 2 H (carbamino-Hb) Transamination eg in metabolism pathway RCHCOOH + RCCOOH RCCOOH + RCCOOH NH 2 O O NH 2 oxidative Deamination eg. in metabolism pathway RCHCOOH RCCOOH + NH 3 NH 2 O
Contributing properties from R groups When R group is plain hydrocarbon (gly, ala, leu, isoleo, val) the a.a interact poorly with water. * When R group have functional groups capable of hydrogen bonding e.g -OH ( Ser, thr, tyr) ; -COOH (asp and glu), these a.a are Hydrophilic or water-loving so easily interact with water. Ester Formation by –OH of serine -OH + H 3 PO 4 phosphoproteins -OH + polysaccharide O-glycoprotein * When R group have functional group –COOH ( asp, glu) the a.a can exist as –ve molecule physiological pH and can form ionic bonds with basic amino acids. When R group have functional group –NH 2 / -NH (lys and hist), these a.a are +ve charged at physiological pH and can form ionic bonds with acidic amino acids. The sulfhydryl group of cysteine is highly reactive. -Oxidation of two molecules of cysteine forms cystine. The 2 molecules is linked by a disulfide bond/bridge. The reaction is reversible oxidation
Transmethylation methyl group of methionine may be transferred to an acceptor to become intermediates in metabolic pathway Formation of S-S bridge. sulfhydryl (-SH) group of cysteine can form the S-S bond with another cysteine residue intrachain or interchain 2 cysteines cystine Function of R groups is also very significant in function of peptides and Proteins. Few examples: a) The hydrophobic aa will generally be found in the interior of proteins shielded from direct contact with water b) The hydrophilic aa will generally be found in the exterior & active centre of enzyme. c) The imidazole ring of histidine acts as proton donor or acceptor at physiological pH hence it is normally found in active site of enzyme, in hemoglobin (RBC).
Few aa are origin/starting molecules for important compounds or amino acid derived molecules: Glutamic acid Gammaaminobutyric acid (GABA) Tyrosine dopamine. these are neurotransmitters. Histidine histamine, a mediator of allergic reactions Tyrosine thyroxine, a thyroid hormone Serine cycloserine an anti-tuberculous; azaserine, an anti-cancer molecule Arginine ornithine and citrulline, intermediates in urea cycle
2. Structure and function of proteins To enable to: Describe the formation of peptide bonds Describe the four levels of protein organization with reference to primary, secondary, tertiary and quaternary structure of proteins using haemoglobin as example Explain how structure of protein determines its function by looking at examples Differentiate between globular and structural proteins with examples eg immunoglobulin, hemoglobin, collagen, keratin etc Describe the functions of protein Relationship between structural protein and its function in health and disease.
Proteins: Biological Functions as biological catalysts of the chemical reactions that occur within the cell examples: i- starch maltose + shorter chain starch ii- protein amino acids + peptide chain iii- triglyceride f f a + mono + di iv- ATP ADP +P i glycerides phosphatase α-amylase trypsin lipase
As regulatory proteins. These proteins regulate the activities of the cell and the ability of other proteins to carry out their cellular function in regulating overall metabolism, growth, development, and maintenance of the organism eg peptide and protein hormones; allosteric enzyme; gene inducers & repressors. As transporter molecules eg. hemoglobin; GLUT,SGLUT i- hemoglobin transport O 2 from tissue to lungs; myoglobin transport O 2 intracellular ii- GLUT transport glucose/galactose from intestinal to blood, iii- SGLUT transport glucose from intestinal to blood. As storage proteins eg myoglobin, stores O 2 in muscle tissue
A peptide bond (amide bond): - feature bonds between amino acids (aa) in polypeptides and proteins. - is formed when the carboxyl group of one aa molecule reacts with the amine group of the other aa molecule in front of it, thereby releasing a molecule of water (H 2 O). - this is a dehydration synthesis reaction or condensation reaction, - the resulting CO-NH bond is called a peptide bond, and the resulting molecule is an amide. The four-atom functional group -C(=O)NH- is called an amide group or (in the context of proteins) a peptide group. - living organisms employ enzymes to form peptide bonds. eg. during translation process. - When two amino acids are linked together, the product is called a dipeptide and when the product is of three amino acids then it is tripeptide
Peptide bond CN O H feature bonds between amino acids (aa) in polypeptides and proteins. is a bond formed when a carboxylic group reacts with an amino group instantaneously eliminating a molecule of H 2 O this is a dehydration synthesis reaction or condensation reaction, the resulting CO-NH bond is called a peptide bond, and the resulting molecule is an amide. The four-atom functional group -C(=O)NH- is called an amide group or (in the context of proteins) a peptide group. living organisms employ enzymes to form peptide bonds. eg. during translation process. When two amino acids are linked together, the product is called a dipeptide and when the product is of three amino acids then it is tripeptide
Structure organization in proteins Primary Structure (or primary level of organization) Definition. Is "The sequence of amino acids in the polypeptide chain.", The N-terminal on the left and C terminal on the right. chain has 50 to 2000 amino acid residues so it is a polypeptide The residues are joined by peptide bonds Changes in the primary structure can alter the proper functioning of the protein.eg offcoded of 2 amino acid in the protein of the glycoprotein in RBC results in MN blood group
N-terminal C-terminal In actual chain these R groups will be the various side chains Peptide bond
At neutral pH Protein with basic aa will have overall positive charge. And that with acidic aa will have overall negative charge Effect of surrounding pH on the structure
cont Effect of surrounding pH on the structure
Secondary structure: There are two types : the α -helix and the β -pleated sheet. The attraction between the R groups can occur within the same chain (case I) or between chains lying next to one another (case II). Case I leads to formation of weak bonds eg hydrogen bonds ; R-R attraction etc. The hydrogen bonds is "Intrachain Hydrogen Bonding" which is between the hydrogen and oxygen atoms of the amino acid backbone. These intrachain weak bondings can cause the chain to twist into a "right handed" coil or α -helix. Case II leads to formation of β -pleated sheet. Such secondary structure α -helix often predominate in "globular proteins and β -pleated sheet predominate in fibrous proteins.
Globular proteins are (i) compactly folded and coiled somewhat spherical. The molecules apolar a.a bound towards the molecule interior and the polar a.a bound towards the molecule exterior allowing dipole-dipole interaction with the solvent. (ii) Soluble in aqueous medium giving colloidal solution (iii) Play numerous functions, as: i) enzymes eg esterases ii) messengers/hormones eg. Insulin iii) transporter of molecules across membran iv) storage eg myoglobin ** α -helix: "alpha" means, looking down the length of the spring, the coiling is happening in a clockwise direction β - pleated sheets: the chains are folded so they lie alongside each other β-pleated, anti-parallel (arrows running in opposite direction H 2 bond
Myoglobin - first globular protein whose structure was analysed by X- ray diffraction by protein crystals. The periodic repeats characteristic of alpha helix were recognised, and the structure shown to have 70% of the polypeptide is alpha- helical. - it is O 2 storage site in muscle tissue. - It is also intracellular transporter of O 2. - It s tertiary (3-D) structure consists of a 8 α-helices which fold to make a compact globular protein. - - the side facing the interior having amino acids with hydrophobic side-chains ie. hydrophobic groups are on the inside of the protein. The side facing to outside having polar side-chains ie. hydrophillic groups are on the outside of the protein, facing the aqueous environment.
Reference: J.Mol. Biol. 142, Myoglobin Structure Heme with Fe 2+/3+ A representation of the 3D structure of the myoglobin protein. Alpha helices are shown in colour, and random coil in white,
β -pleated sheet - the β -pleated sheet forms when the hydrogen atoms of the amino group and the oxygen atoms of the carboxyl group of amino acids on two chains (or more) lying side-by-side forms hydrogen bond. - Closely associate to structural/fibrous proteins - the protein chains are in associate to form long fibers - elongated or needle shaped - possess minimum solubility - - resist digestion - The β -pleated sheet structure is often found in many structural proteins, eg "Fibroin", the protein in spider webs; Keratin- a structural protein found in hair and nails, skin, and tortoise shells
Fibrous proteins are more filamentous or elongated, play only structural funtions. Also known as scleroproteins. Found only in animals. Are water-insoluble. Used to construct connective tissues, tendons, bone matrix, muscle fibers. Examples are keratin (hair; tough and hard bud not mineralized structure as in reptiles), collagen ( long chains, tied into bundles, has great tensile strength). Its degradation leads to wrinkles that accompanying aging.
"Tertiary" Structure: a 3 dimensional chain arrangement, the way the whole chain (including the secondary structures) folds itself into its final 3-dimensional shape. is held together by interactions between the side chains - the "R" groups. Interactions such as: ionic; van der Waals (hydrophobic-hydrophobic); H-bonds; S-S bridge OR When "proline", an oddly shaped amino acid occurs in the polypeptide chain a "kink" in the a-helix develops. Kinks can also be caused by repulsive forces between adjacent charged R groups. These kinks create a 3 dimensional chain arrangement. This 3 dimensional shape is also held together by weak hydrogen bonds "disulfide" bonds between two amino acids of cystine ("covalent") disulfide "bridges" (linkages) cystine -- s -- s – cystine. These strong covalent bonds hold the protein in its specific 3D shape. The 3D shape creates "pockets" or "holes' in the surface of the protein which are very important in enzyme function
pleated sheets random coils α-helix Cystinyl
Quaternary Structure of Proteins 2 or more 3 dimensional tertiary proteins and sticking them together to form a larger protein. Many enzymes and transport proteins are made of two or more parts. only exists, if there is more than one polypeptide chain present in a complex protein Hemoglobin: an oxygen carrying protein in red blood cells which is made of 4 parts.
Structural Level of Proteins
Denaturation or Loss of 3-D shape denaturing agents: Temperature> 40 o C; mineral acids; salts. eg. when heated, protein can unfold or "Denature". This loss of three dimensional shape will usually be accompanied by a loss of the proteins function. If the denatured protein is allowed to cool it will usually refold back into its original conformation.
Protein metabolism denotes the various processes responsible for the denotes the various processes responsible for the (i) biosynthesis of proteins from amino acids. (i) biosynthesis of proteins from amino acids. (ii) catabolism the breakdown of proteins by /proteolysis liberating of amino acids. (ii) catabolism the breakdown of proteins by /proteolysis liberating of amino acids. That is, comprises of I- Protein metabolism (synthesis and breakdown) II-Amino Acid metabolism (synthesis and breakdown) WILL PROCEED WITH Protein metabolism (synthesis and breakdown) Protein metabolism (synthesis and breakdown)
PROTEIN SYNTHESIS proteins of one organ are similar but differ from that of another organ. That is, each chain is characterized by a specific sequence of a.a. How is this special feature achieved? proteins of one organ are similar but differ from that of another organ. That is, each chain is characterized by a specific sequence of a.a. How is this special feature achieved? The sequence of a.a in a particular chain is ensured through the following units and process: The sequence of a.a in a particular chain is ensured through the following units and process: translation; Codons; transciption tRNA; mRNA; translation; Codons; transciption tRNA; mRNA;
Translation is process of protein synthesis. It is translating genetic messages into the primary sequence of a polypeptide. tRNA carries a specific amino acid to the matching position along the mRNA template. It can be divided into 4 stages: Activation, Initiation, Elongation and Termination, each regulated by a large number of proteins and coactivators. It occurs in cytoplasm. Translation is process of protein synthesis. It is translating genetic messages into the primary sequence of a polypeptide. tRNA carries a specific amino acid to the matching position along the mRNA template. It can be divided into 4 stages: Activation, Initiation, Elongation and Termination, each regulated by a large number of proteins and coactivators. It occurs in cytoplasm. Codon: a sequence of 3 nucleotide in DNA that codes a single a.a Codon: a sequence of 3 nucleotide in DNA that codes a single a.a transcription : synthesis of a single strand messenger RNA (mRNA) by transcribing the sequence of the nucleotide in the template DNA/genom. The reaction is catalyzed by RNA polymerase. The template DNA is unzipped by enzyme helicase prior to the transcription. transcription : synthesis of a single strand messenger RNA (mRNA) by transcribing the sequence of the nucleotide in the template DNA/genom. The reaction is catalyzed by RNA polymerase. The template DNA is unzipped by enzyme helicase prior to the transcription.
tRNA is transfer RNA that carries an a.a to the mRNA to be incorporated into the peptide chain. tRNA is transfer RNA that carries an a.a to the mRNA to be incorporated into the peptide chain. mRNA is a type of RNA that encoding the sequence of the protein in the form of a trinucleotide code. The specific sequence of the nucleotide is accomplished through transcription. mRNA is a type of RNA that encoding the sequence of the protein in the form of a trinucleotide code. The specific sequence of the nucleotide is accomplished through transcription. trinucleotide code trinucleotide code
Activation: the correct amino acid (AA) is joined to the correct tRNA. The AA is joined by its carboxyl group to the 3' OH of the tRNA by an ester bond. The anti-codon determines the correct AA. Activation: the correct amino acid (AA) is joined to the correct tRNA. The AA is joined by its carboxyl group to the 3' OH of the tRNA by an ester bond. The anti-codon determines the correct AA. Initiation: involves the small subunit of the ribosome binding to 5' end of mRNA with the help of initiation factor (IF), Initiation: involves the small subunit of the ribosome binding to 5' end of mRNA with the help of initiation factor (IF), Elongation occurs when the next aminoacyl-tRNA (charged tRNA) in line binds to the ribosome along with GTP and an elongation factor. Elongation occurs when the next aminoacyl-tRNA (charged tRNA) in line binds to the ribosome along with GTP and an elongation factor. Termination of the polypeptide happens when the A site of the ribosome faces a stop codon (UAA, UAG, or UGA).This activates release factor which then causes the release of the polypeptide chain. Termination of the polypeptide happens when the A site of the ribosome faces a stop codon (UAA, UAG, or UGA).This activates release factor which then causes the release of the polypeptide chain. The synthesis process/translation
TRANSLATION in diagrame : LOADED tRNA RIBOSOME mRNA COMPONENTS PRESENT IN THE PROCESS anticodon Aminoacid carried codon
TRANSLATION The newly made mRNA (transcription) leaves the nuceus and binds with the ribosome in the cytoplasm. ONE codon is exposed at site P and another codon at site A A tRNA with a complementary codon in its anticodon site will bind with the codon at site P, bringing an aminoacid. 1º AMINOACID: Methionine (AUG) in site P.
TRANSLATION Even though every protein begins with the Methionine amino acid, not all proteins will ultimately have methionine at one end. If the "start" methionine is not needed, it is removed before the new protein goes to work (either inside the cell or outside the cell, depending on the type of protein synthesized)
TRANSLATION A 2º AMINOACID: Glycine (only in this case) in site A. PEPTIDIC BOND IS FORMED
TRANSLATION STOP codon NO aminoacid is added. Its the END of the polypeptide! Growing polypeptide
PROTEIN CATABOLISM Has various indication: Comprises of Digestion and Absorption Is carried out via proteolysis is the directed degradation (digestion) of proteins by cellular enzymes called proteases (various kinds) releasing peptide/A.Aproteinsenzymesproteases The digestion of proteins from foods as a source of amino acids (aas)amino acids The aas constituting aa pool are metabolized further ( aa catabolism)
Digestion: Source of proteins that come in the diet: - animal eg milk, dairy products, meat, fish, eggs, liver - vegetable sources eg cereals, pulses, peas, nuts and beans In mouth: no proteolytic enzyme so the proteins are unchanged but the size(food) becomes smaller due to mastication and chewing. Food bolus travels down and reaches stomach and meet gastric juice In stomach ( pH 1-2 maintains by HCl) : attack by pepsin, renin, gelatinase and gastricin ( enzymes in the gastric juice). All these enzymes attack internal peptide bonds. - Pepsin( a endoproteinase) acts on : Proteins proteoses + peptones Casein(milk) paracasein + proteos (whey proteins) paracasein + Ca 2+ calcium paracasein (insoluble) - gastricin ( a proteinase) - gelatinase: gelatine polypeptide DIGESTION & ABSORPTION
In small intestine : duodenum, jejunum, ileum - Duodenum: Food bolus meet pancreatic juices. Enzymes in pancreatic juices : Trypsin ( a proteolytic enzyme) Chymortypsin Carboxy peptidases ( 2 types: A and B) are exopeptidases; splits one amino acid at a time fr free end. Elastases : a serine protease Collagenases act on protein present in collagen/connective tissue yielding peptide - Jejunum-ileum Food remnant meet intestinal juice. Enzymes in intestinal juices: Amino peptidase: peptides tripeptides Enteropeptidase/Enterokinase Prolidase: acts at terminal proline Di and tri-peptidase: Di and tri-peptide amino acids
Absorption Of Amino Acids Absorption is by active transport Site of absorption is - ileum and distal jejunum: amino acids - duodenum and proximal jejunum: di and tri-peptides After absorption, amino acids and di and tri-peptides (if any) are carried by portal blood to liver, partly : i- are taken up by liver cells ii- enter the systemic circulation (made up part of aa pool), diffusing throughout body fluid & taken up by tissue cells. ( ( The body's circulatory system has three distinct parts: pulmonary (the lungs) circulation, coronary (the heart) circulation, and systemic (the rest of the system) circulation. Each part works independently in order for them to all work together)
The aa will be used to synthesize: tissue proteins; enzyme; hormones 3 states relates to aa pool -cell : i- dynamic equilibrium amnt of aa taken-up = amnt of aa loss ii- cell waste amnt of aa taken-up < amnt of aa loss iii- cell grows amnt of aa taken-up > amnt of aa loss Regulatory of Amino Acid If amino acids are in excess of the body's biological requirements, they are metabolized to glycogen or fat and subsequently used for energy metabolism. If amino acids are to be used for energy their carbon skeletons are converted to acetyl CoA, which enters the Krebs cycle for oxidation, producing ATP
Summary: Digestion & Absorption Aas available for use in metabolic processes come from dietary protein and breakdown of tissue protein by proteolysis. Digestion (dietary protein) occurs in stomach as well as intestine. In stomach, digested by pepsin, in intestine and duodenum by a group of enzymes, protease (trypsin. Chymotrypsin and carboxypeptidase) These liberated aas are absorbed into cells and are collectively referred as aa pool ## Amino acids are transported into cell by various transport mechanisms involving membrane-bound transport proteins. Ingested protein digested aa absorbed (aa pool)
Dietary protein Tissue protein enzymic proteolysis e.g trypsin/pepsin A A pool NH 2 C skeleton Excreted as urea & uric acid protein energy Precursors for other molecules synthesis New AA N containing molecules Healthy & in young subject >> aa breakdown INSIDE CELL OUTSIDE CELL Assimilation of Amino Acids
Amino acid synthesis is the set of metabolic pathways /processes by which the various amino acids are produced from direct incorporation/ combination (I) of –NH 2 group OR (II) of ammonium ion NH 4 + with other compounds found in the organisms diet or growth media I –NH 2 group is incorporated into α-keto acid through 2 types of reactions: i- non-reductive transamination i) glutamate/aspartate as –NH 2 donor ii) glutamine/asparagine as –NH 2 donor iii) branch chain aa as –NH 2 donor ii- reductive transamination II- ammonium ion NH 4 + is incorporated α-keto acid. through - Reductive amination - non-reductive amination
Non-reductive transamination characteristics:- Non-reductive transamination characteristics:- Reaction of Glutamate (or Aspartate) and an α-keto acid or BCAA. -NH 2 is transferred from Glutamate/ Aspartate to an α-keto acid. ( Glutamate/ Aspartate/asparagine is -NH 2 group donor; α-keto acid supplies C-skeleton) ** glutamate as -NH 2 group donor is more regular Reaction is catalysed by i) enzyme aminotransferase or transaminase ; ii) required co-enzyme pyridoxal-5-phosphate (PLP) R1CC--O + R1CHC--O + α-ketoglutarate O O + NH 3 New AA Acceptor α-ketoacid referred as pair O
i) Glutamate and an α-keto acid (pyruvate) i) Glutamate and an α-keto acid (pyruvate) ii) Aspartate and an α-keto acid ii) Aspartate and an α-keto acid (pyruvate) NH 2 CH 3 CH 3 OO C-CH 2 -CH 2 - CH + C=O HC NH 2 + OOC-CH 2 -CH 2 C=O COOH COOH COOH COOH Glutamate pyruvate alanine α-ketoglutarate Glutamate pyruvate alanine α-ketoglutarate NH 2 CH 3 CH 3 OO C-CH 2 -- CH + C=O HC NH 2 + OOC--CH 2 C=O COOH COOH COOH COOH aspartate pyruvate alanine oxaloacetate aspartate pyruvate alanine oxaloacetate pair Non-reductive transamination: Examples
Non-reductive transamination (in skeletal muscle). Non-reductive transamination (in skeletal muscle). enzyme: glutamine synthase (GS) enzyme: glutamine synthase (GS) Glutamate + BCAA glutamine + α-keto acid Glutamate + BCAA glutamine + α-keto acid ( BCAAs are comprised of valine, leucine, and isoleucine) ( BCAAs are comprised of valine, leucine, and isoleucine) OOC-CH 2 -CH 2 -CH + (CH 3 ) 2 CH-CH O=C-CH 2 -CH 2 -CH + (CH 3 ) 2 -CH-C NH 2 NH 2 NH 2 NH 2 O COO Glutamate valine glutamine BC α-oxoacid
Enzyme Transaminase/Aminotransferase Enzyme Transaminase/Aminotransferase requires co-enzyme pyridoxal-5-PO 4, abbreviated (PLP). requires co-enzyme pyridoxal-5-PO 4, abbreviated (PLP). a derivative of vitamin B 6 a derivative of vitamin B 6 R of Lysine
PLP attaches to the active site of enzyme by noncovalent interaction and a Schiff base aldimine ( condensation of ε- amino of lysine residue and aldehyde group of PLP) is formed. amino acid substrate becomes bound to PLP via the α- amino group in an imine exchange reaction. bond 1 breaks leaving –NH 2 on the co-enzyme to be transferred to an α-keto acid, [ Vitamin B6 is involved in the metabolism (especially catabolism) of amino acids, as a cofactor in transamination reactions. This is the last step in the synthesis of nonessential amino acids and the first step in amino acid catabolism. Vitamin B6 is a mixture of pyridoxin derivatives. PLP is 1 of them].
Reductive Transamination Glutamine, asparagine transfer the amide nitrogen to oxo (or keto) acid to form a new amino acid. 2-oxoglutarate is –NH 2 receptor and glutamine is –NH 2 donor The enzyme GOGAT is NADPH dependent glutamine + 2-oxoglutarate + NADPH + H + ---> 2 glutamate + NADP + GOGAT: enzyme glutamine oxoglutarate amidotransferase GOGAT
i) Reductive amination i) Reductive amination reaction of α-ketoglutarate with NH 4 + leading to formation of glutamate (in mitochondria & cytoplasm). reaction of α-ketoglutarate with NH 4 + leading to formation of glutamate (in mitochondria & cytoplasm). α-ketoglutarate is –NH 2 acceptor α-ketoglutarate is –NH 2 acceptor catalysed by glutamate dehydrogenase, the enzyme is NADH dependent catalysed by glutamate dehydrogenase, the enzyme is NADH dependent reaction is reversible i.e the reverse pathway is a primary means of producing NH 4 + for N excretion. reaction is reversible i.e the reverse pathway is a primary means of producing NH 4 + for N excretion. The enzyme is driven toward right when excess NH 4 + is present The enzyme is driven toward right when excess NH 4 + is present NH 4 + is from oxidative deamination of glutamate (in extrahepatic tissue) NH 4 + is from oxidative deamination of glutamate (in extrahepatic tissue) II- Incoporation of NH 4 + ion:
+ NH NADH + H + Enzyme: Glutamate Dehydrogenase + NAD + + H 2 O Reductive Amination : left - right (Oxidative Deamination : right left) GD NH 3 + H +
ii) Non-reductive amination or amidation Glutamate or aspartate react with NH 4 + to form glutamine, (asparagine) Glutamate or aspartate react with NH 4 + to form glutamine, (asparagine) catalyze by glutamine/ asparagine synthetase respectively. catalyze by glutamine/ asparagine synthetase respectively. Sites : liver, brain, kidney, muscles & intestine Sites : liver, brain, kidney, muscles & intestine This rxn forms the path by which cell rid off excess NH 4 +. This rxn forms the path by which cell rid off excess NH 4 +. ** NH 4 + at high conc may be toxic to certain cell e.g brain cell. Glutamine is non toxic. ** NH 4 + at high conc may be toxic to certain cell e.g brain cell. Glutamine is non toxic.
COO-CH 2 -CH 2 - CH NH 3 + COO - + ATP + NH 4 + CO-CH 2 -CH 2 - CH NH 2 NH 3 + COO - Glutamine + ADP Glutamine Synthetase (GS) + Pi Glutamate COO-CH 2 - CH From excess aa pool COO - NH ATP + NH 4 + Asparagine Synthetase CO-CH 2 -CH NH 2 NH 3 + COO - Asparagine + ADP + Pi Aspartate Non-reductive amination or amidation
Glutamine synthetase (GS) catalyzes the ligation of glutamate and ammonia to form glutamine, with concomitant hydrolysis of ATP. In mammals, the activity eliminates cytotoxic ammonia, at the same time converting neurotoxic glutamate to harmless glutamine; there are a number of links between changes in GS activity and neurodegenerative disorders, such as Alzheimer`s disease.
glutamate α-ketoglutarate NH 4 + GD NADH glutamate Oxidative deamination Reductive amination α-keto acid transamination New aa
C skeleton of all non-essential aa are derivatives of: C skeleton of all non-essential aa are derivatives of: Glycerate -3-phosphate Glycerate -3-phosphate Pyruvate Pyruvate Α-ketogluterate Α-ketogluterate Oxaloacetate Oxaloacetate But Tyrosine from essential aa phenylalanine But Tyrosine from essential aa phenylalanine On basis of common precursor Ξ similarities in their synthetic On basis of common precursor Ξ similarities in their synthetic Pathway, aa can be grouped into 5 families.
glutamate family= synthesis of glutamate, glutamine, arg, pro. glutamate family= synthesis of glutamate, glutamine, arg, pro. - C skeleton derive fr α-ketoglutarate - C skeleton derive fr α-ketoglutarate serine family = synthesis of serine, glycine, cystein serine family = synthesis of serine, glycine, cystein - C skeleton derive fr glycerate-3-phosphate - C skeleton derive fr glycerate-3-phosphate aspartate family = synthesis of aspartate, lysine, methionine, asparagine, threonine aspartate family = synthesis of aspartate, lysine, methionine, asparagine, threonine - C skeleton derive fr oxaloacetate - C skeleton derive fr oxaloacetate pyruvate family = synthesis of alanine, valine, leucine, isoleucine pyruvate family = synthesis of alanine, valine, leucine, isoleucine - C skeleton derive fr pyruvate - C skeleton derive fr pyruvate aromatic family = synthesis of *phenylalanine, tyrosine, *tryptophan *EAA aromatic family = synthesis of *phenylalanine, tyrosine, *tryptophan *EAA
Glutamate family - key substrate is α- ketoglutarate fr TCA -Glutamate is produced by GD and is the principle rxn of fixation of NH 3 in bactria - glutamine is produced by ATP-requiring +n of NH 3 to glu and the rxn fnc as a major means of assimilating of NH3 fr environment -Regulation of this family is controlled by repression of mRNA and feedback inhibition: by prolin and arg
Regulatory of Amino Acid If amino acids are in excess of the body's biological requirements, they are metabolized to glycogen or fat. If amino acids are in excess of the body's biological requirements, they are metabolized to glycogen or fat. If amino acids are to be used for energy their carbon skeletons are converted to acetyl CoA, or other metabolites intermediates (pyruvate, oxaloacetate, Succinyl-coA ) which then enters the Krebs cycle for oxidation, producing ATP. If amino acids are to be used for energy their carbon skeletons are converted to acetyl CoA, or other metabolites intermediates (pyruvate, oxaloacetate, Succinyl-coA ) which then enters the Krebs cycle for oxidation, producing ATP.
Catabolism of AA Generally involves : Generally involves : - Removal of amino group - Disposal of amino group to final compounds urea([NH 2 ] 2 CO) and ammonia (NH 3 ); also incoporated into other molecules - Utilization of C skeleton by channeling into TCA through which they are converted to final products carbon dioxide (CO 2 ), water (H 2 O), ATP, or degraded into a variety of metabolite intermediates which then enter synthesis pathway of other compounds or degraded into a variety of metabolite intermediates which then enter synthesis pathway of other compounds - Decarboxylation - one carbon metabolism
Removal of amino group Removal of amino group Occurs by Occurs by - transamination - transamination - oxidative deamination (only happens with glutamate ) catalyses by glutamate dehydrogenase - oxidative deamination (only happens with glutamate ) catalyses by glutamate dehydrogenase glutamate + NAD + NH α-ketoglutarate glutamate + NAD + NH α-ketoglutarate Transamination ( largely occurs in cytosol of liver cells) is the transfer of the nitrogen (the amino) group of an L-a.a to α-ketoglutarate forming L-glutamate. The reaction is catalysed by transaminase and it requires co-enzyme pyridoxal-5-PO 4 (see earlier section for detail mechanism). Glutamate may undergo another transamination, transfering –NH 2 to another α-ketoacid i.e glutamate becomes -NH 2 carrier
Oxidative Deamination (O.xdn) reaction is prevalent when protein intake > protein synthesis => aa fromaa pool undergoes degradation. The N- in aa is removed by deamination rxn and converted to ammonia which is toxic, therefore need to be detoxified and excreted. Is :L-glutamate + NAD + NH α-ketoglutarate happens only with glutamate catalyses by glutamate dehydrogenase GD. It occurs in liver & in most extrahepatic tissue.
* N of amino group made available for excretion by rxn. In muscle cell ( no GD) any excess aa transfer its -NH 2 to α - ketoglutarate to form L-glutamate (transamination). L- glutamate undergoes transamination with pyruvate catalyse by alanine transaminase to give alanine + α -ketoglutarate. Alanine carries by blood to liver, (alanine cycle). In liver, alanine + α -ketoglutarate react catalysed by alanine transaminase reforming L-glutamate + pyruvate as alanine transaminase rxn is reversible. Then L-glutamate undergoes Oxidative deamination. Pyruvate can be diverted to gluconeogenesis. This process is refered to as the glucose-alanine cycle and NH + 4 moves onto urea cycle which is also known as ornithine cycle, be converted to urea. Urea is transferred through the blood to the kidneys and excreted in the form of urine.
glutamate α-ketoglutarate NH 4 + Alanine transamination H 2 O + NAD + Transported to liver for Oxidative deamination Alanine transamination New α-keto acid transamination excess aa pyruvate alanine -NH 2 in Muscle NH 4 + (liver) Liver alanine α-ketoglutarate glutamate pyruvate GD α-ketoglutarate + NADH Alanine Cycle To urea cycle
Deamination is also an oxidative reaction occurs under aerobic conditions in all tissues but especially the liver. During oxidative deamination, an amino acid is converted into the corresponding keto acid by the removal of the amine functional group as ammonia and the amine functional group is replaced by the ketone group. The reaction is catalysed by glutamate dehydrogenase which is allosterically controlled by ATP and ADP. ATP acts as an inhibitor whereas ADP is an activator. The ammonia eventually goes into the urea cycle. Oxidative deamination occurs primarily on glutamic acid because glutamic acid was the end product of many transamination reactions.
The glutamate dehydrogenase (GD) is allosterically controlled by ATP and ADP. ATP acts as an inhibitor whereas ADP is an activator. GD
Summary of Urea Cycle Summary of Urea Cycle Occurs in liver cells Occurs in liver cells Is a 5 steps cycle: 1 step in mitochondria 4 steps in cytosol Is a 5 steps cycle: 1 step in mitochondria 4 steps in cytosol Main substrates: NH 3, CO 2 and Aspartate. Main substrates: NH 3, CO 2 and Aspartate. In the matrix of mitochondria occurs CPS I and OTC catalysed rxn, In the matrix of mitochondria occurs CPS I and OTC catalysed rxn, CPS rxn uses 2ATP and reaction is irreversible CPS rxn uses 2ATP and reaction is irreversible Citrulline Ornithine occur in cytosol, in 4 steps Citrulline Ornithine occur in cytosol, in 4 steps -Citrulline is tranported across the inner membrane -Citrulline is tranported across the inner membrane by a carrier neutral aa. - enzymes are arginosuccinate synthase, arginosuccinate lyase and arginase - enzymes are arginosuccinate synthase, arginosuccinate lyase and arginase Urea transferred to kidney through blood and excreted as urine Urea transferred to kidney through blood and excreted as urine
Fate of A.A Nitrogen Excreted in the form of urea (urine) Transferred to specific α-keto acids (of the TCA intermediates) to form new a.a. This can be represented in the form of α-keto acids / aa pair eg: α-ketoglutarate/glutamate; pyruvate/alanine; aspartate/oxaloacetate pair. Incorporated into skeleton of non amino acid molecules => aa derived compound
Derived AA Compounds What are derived amino acid compounds? They are compounds that contain N- atom, S- atom or part of aa structure as part of their molecular structure.Can be divided into 2 groups: alkaloids (in plants) & animal related. Animal related and specific parent aa eg. Glutathione (GSH), Serotonin and Histamine, Heme, GABA, DNA bases Why the synthesis occurs? These molecules are synthesized because they are important to the body. The synthesis process
serotonin Fncn to influence the functioning of the cardiovascular, renal, immune, and gastrointestinal systems Fncn to influence the functioning of the cardiovascular, renal, immune, and gastrointestinal systems Any disruption in the synthesis, metabolism or uptake of this neurotransmitter has been found to be partly responsible for certain manifestations of schizophrenia, depression, compulsive disorders and learning problems. Any disruption in the synthesis, metabolism or uptake of this neurotransmitter has been found to be partly responsible for certain manifestations of schizophrenia, depression, compulsive disorders and learning problems.schizophrenia depressionschizophrenia depression Synthesis: Synthesis:
Function of some AA derived compounds As neurotransmitter : GABA, dopamine, serotonin, As neurotransmitter : GABA, dopamine, serotonin, Sleep inducing : melatonin Sleep inducing : melatonin Carrier : carnithine Carrier : carnithine As hormone: tyroxine, As hormone: tyroxine, Dilating/constriction of blood vessel: histamine Dilating/constriction of blood vessel: histamine Exhibit multifunctions: GSH Exhibit multifunctions: GSH - acts as reducing agent in NA and eicosanoids synthesis - acts as reducing agent in NA and eicosanoids synthesis - maintain the sulfahydryl grp of enzymes & other molecules in reduced state - maintain the sulfahydryl grp of enzymes & other molecules in reduced state - promotes aa transport - promotes aa transport - protect cells fr radiation, O 2 toxicity and environmental toxins - protect cells fr radiation, O 2 toxicity and environmental toxins
Utilization of the C-skeleton The C-skeleton of the standard amino acids are degraded to seven common metabolic intermediates such as Acetyl-coA; Acetoacetyl-CoA; pyruvate; Oxaloacetate, α-ketoglutarate, Succinyl-CoA and fumerate. Succinyl-CoA and fumerate. Those aa are referred to different names depending to the class to which the final product are classified: i) degraded to acetyl-CoA and AceAcetyl-CoA are referred to as KETOGENIC because the intermediates lead to either fatty acids or ketone bodies.eg Lys and Leu i) degraded to acetyl-CoA and AceAcetyl-CoA are referred to as KETOGENIC because the intermediates lead to either fatty acids or ketone bodies.eg Lys and Leu
ii) degraded to pyruvate; α-ketoglutarate, Succinyl- CoA, Oxaloacetate, and fumerate are referred to as GLUCOGENIC because they are intermediates of gluconeogenesis. All except Lys and Leu are pure or partly glucogenic Those that yield acetyl-CoA are divided into 2 groups. Those that yield acetyl-CoA are divided into 2 groups. a) Those that yield pyruvate as intermediate: Ala, Cys, Gly, Ser and Thr a) Those that yield pyruvate as intermediate: Ala, Cys, Gly, Ser and Thr b) Those that do not yield pyruvate as intermediate: Phe, Lys, Leu Trp and Tyr b) Those that do not yield pyruvate as intermediate: Phe, Lys, Leu Trp and Tyr
utilization of the C-skeleton
Decarboxylation of amino acid is effected by decarboxylase enzyme, PLP dependent Products are alkylamine + CO 2. The alkylamine are neurotransmitters There are 4 aa decarboxylase enzymes: Aromatic L-amono acid decarboxylase (is a group of enzymes); L-glutamate decarboxylase (GAD); lysine decarboxylase (LDC); histidine decarboxylase (HDC)
HDC HOOC-CH 2 -CH 2 -CH(NH 2 )-COOH CO 2 + HOOC-CH 2 -CH 2 -CH 2 NH 2 GABA is a neurotransmitter in brain GAD (GABA) Aromatic L-aa decarboxylase synonyms to DOPA decarboxylase, tryptophan decarboxylase, 5- hydroxytryptophan decarboxylase, AAAD.tryptophandecarboxylase tryptophan tryptamine + CO 2 Tryp D
A.As Metabolic Disorder Diseases - Are diseases resulted from disorders of a.as processing/metabolism due to Inherited/genetic defects that cause deficiency of certain enzymes for Inherited/genetic defects that cause deficiency of certain enzymes for i) the breakdown of amino acids or i) the breakdown of amino acids or ii) the body's ability to get the amino acids into cells or ii) the body's ability to get the amino acids into cells or iii) Amino acid Transport iii) Amino acid Transport - Symptoms of disease appear early in life - Generally are autosomal recessive that is why only small number of man suffers.
Inherited metabolic disorder ( I.M.D) : Inherited metabolic disorder ( I.M.D) : Oculocutaneous albinism Oculocutaneous albinism Tyrosinemia of tyrosine Tyrosinemia of tyrosine Alkaptonuria Alkaptonuria Phenylketonuria of phenylalanine Phenylketonuria of phenylalanine Hyperalaninemia Hyperalaninemia Leucinosis or maple syrup urine disease – of branched-chain a.a Leucinosis or maple syrup urine disease – of branched-chain a.a homocystinuria – of methionine homocystinuria – of methionine Nonketotic hyperglycinemia – of glycine Nonketotic hyperglycinemia – of glycine
PROTEIN CATABOLISM Has various indication: Has various indication: Is carried out via proteolysis Is carried out via proteolysis is the directed degradation (digestion) of proteins by cellular enzymes called proteases (various kinds) releasing peptide/A.A is the directed degradation (digestion) of proteins by cellular enzymes called proteases (various kinds) releasing peptide/A.Aproteinsenzymesproteasesproteinsenzymesproteases The digestion of proteins from foods as a source of amino acids (aas) The digestion of proteins from foods as a source of amino acids (aas)amino acidsamino acids The aas constituting aa pool are metabolized further (refer to aa catabolism) The aas constituting aa pool are metabolized further (refer to aa catabolism)