Lecture 5 Protein Structure Biochemistry Lecture 5 Protein Structure
Structure of Proteins
Definition of Structural States Primary Structure (1°) – peptide bond linking amino acid residues in a polypeptide chain (sequence of amino acid residues). Secondary Structure (2°) – local spatial arrangement of the main-chain atoms in a selected segment of a polypeptide chain. Tertiary Structure (3°) – overall three-dimensional arrangement of all atoms in a protein. Quaternary Structure (4°) – arrangement of protein subunits (separate polypeptide chains) in three-dimensional complexes.
Chapter 6, Figure 6.1, Three-dimensional folding of the protein myoglobin.
Structural Groups
Chapter 6, Figure 6.2, The four levels of structural organization of proteins
Chapter 6, Figure 6.9, A Ramachandran plot of poly-L-alanine
Keratin
Collagen
Silk
Globular Proteins
2° Structural Motifs (Supersecondary Structures)
Various Interactions Create 3D Shapes
Chapter 6, Figure 6.1, Three-dimensional folding of the protein myoglobin.
Quaternary Structure
X-Ray Crystallography
Nuclear Magnetic Resonance
Chapter 6, Figure 6A.9, Nuclear magnetic resonance spectroscopy (NMR)
How Do We Obtain Proteins for Structural & Functional Analysis?
Purification of Proteins FIGURE 3-16 Column chromatography. The standard elements of a chromatographic column include a solid, porous material (matrix) supported inside a column, generally made of plastic or glass. A solution, the mobile phase, flows through the matrix, the stationary phase. The solution that passes out of the column at the bottom (the effluent) is constantly replaced by solution supplied from a reservoir at the top. The protein solution to be separated is layered on top of the column and allowed to percolate into the solid matrix. Additional solution is added on top. The protein solution forms a band within the mobile phase that is initially the depth of the protein solution applied to the column. As proteins migrate through the column, they are retarded to different degrees by their different interactions with the matrix material. The overall protein band thus widens as it moves through the column. Individual types of proteins (such as A, B, and C, shown in blue, red, and green) gradually separate from each other, forming bands within the broader protein band. Separation improves (i.e., resolution increases) as the length of the column increases. However, each individual protein band also broadens with time due to diffusional spreading, a process that decreases resolution. In this example, protein A is well separated from B and C, but diffusional spreading prevents complete separation of B and C under these conditions. 20
Chapter 5, Figure 5A.6, An overview of ion exchange chromatography
Chapter 5, Figure 5A.7, An overview of size exclusion chromatography
Chapter 5, Figure 5A.3, An overview of affinity chromatography
Protein Analysis - Electrophoresis + + - Heat - - - - - - -
Folding and Denaturation
Chapter 6, Figure 6.24, Contribution to the free energy of folding of globular proteins
Denaturation… also known as “Cooking”
Misfolding Diseases
Mutation Impairs Proper Folding Sickle Cell Anemia Cystic Fibrosis
Contagious Misfolding: Prions