CHAPTER 8 Metabolic Respiration Overview of Regulation Most genes encode proteins, and most proteins are enzymes. The expression of such a gene can be regulated by controlling the activity of the enzyme or controlling the amount of enzyme produced.

An Overview of Gene Regulation

Regulation of Enzyme Activity Noncovalent Enzyme Inhibition Many metabolic reactions can be regulated through control of the activities of the enzymes that catalyze them. An important type of regulation of enzyme activity is feedback inhibition (Figure 8.2), in which the final product of a biosynthetic pathway inhibits the first enzyme unique to that pathway.

Feedback Inhibition

An allosteric enzyme has two binding sites, the active site, where the substrate binds, and the allosteric site, where the inhibitor (called an effector) binds reversibly (Figure 8.3).

Some biosynthetic pathways under feedback inhibition employ isoenzymes, different proteins that catalyze the same reaction but are subject to different regulatory controls (Figure 8.5).

3-deoxy-D-arabinoheptulosonate 7- phosphate Isoenzyme and feedback inhibition

Covalent Modification of Enzymes Covalent modification is a regulatory mechanism for changing the activity of an enzyme. Enzymes regulated in this way can be reversibly modified. One type of modification is adenylylation (the addition of AMP) (Figure 8.6). Others are phosphorylation and methylations

Regulation of glutamine synthetase by adenylation 1. GS inhibition by several a. a. and compounds involved in nucleotide metabolism 2. GS (12 identical subunits) is also inhibited by covalent modification - adenylation Nitrogen rich medium (Lot of glutamine available) Nitrogen poor medium

Protein splicing (Figure 8.7) is a form of posttranslational modification. Topoisomerase II (GyrA) in Mycobacterium leprae (Lyprosae disease)

DNA-Binding Proteins and Regulation of Transcription by Negative And Positive Control Certain proteins can bind to DNA because of interactions between specific domains of the proteins and specific regions of the DNA molecule (Figure 8.8).

DNA Binding Proteins homodimers

In most cases, the interactions are sequence- specific. Proteins that bind to nucleic acid may be enzymes that use nucleic acid as substrates, or they may be regulatory proteins that affect gene expression.

Several classes of protein domains are critical for proper binding of these proteins to DNA. One of these is called the helix-turn- helix motif (Figure 8.9).

Lac and trp repressors and lambda repressor (>250)

Another domain is the zinc finger, a protein that binds a zinc ion (eukaryotes). Cysteine (C) and histidine (H)

Negative Control of Transcription: Repression and Induction

The amount of an enzyme in the cell can be controlled by decreasing (repression, Figures 8.11, 8.13) or increasing (induction, Figure 8.12) the amount of mRNA that encodes the enzyme.

(Arginine) Expression of arginine biosynthetic pathway (Operon)

This transcriptional regulation involves allosteric regulatory proteins that bind to DNA. For negative control of transcription, the regulatory molecule is called a repressor protein and it functions by inhibiting mRNA synthesis.

Positive Control of Transcription Positive control of transcription is implemented by regulators called activator proteins. They bind to activator-binding sites on the DNA and stimulate transcription. As in repressors, activator protein activity is modified by effectors.

For positive control of enzyme induction, the effector promotes the binding of the activator protein and thus stimulates transcription (Figures 8.14, 8.15).

Induction of lac operon (negative regulation) (beta galactosidase)

Maltose operon regulation (positive) (Absence of inducer - maltose) (maltose)

Global Regulatory Mechanisms Global Control and the lac Operon

Global control systems regulate the expression of many genes simultaneously. Catabolite repression is a global control system, and it helps cells make the most efficient use of carbon sources.

The lac operon is under the control of catabolite repression as well as its own specific negative regulatory system (Figure 8.20).

cAMP and CAP(catabolite activator protein)

The Stringent Response The stringent response (Figure 8.21) is a global control mechanism triggered by amino acid starvation. The alarmones ppGpp and pppGpp are produced by RelA, a protein that monitors ribosome activity.

ppGpp

The stringent response achieves balance within the cell between protein production and protein requirements.

Other Global Control Networks Cells can control several regulons by employing alternative sigma factors. Alternative signal factors in Escherichia coli are shown in Table 8.2.

These recognize only certain promoters and thus allow transcription of a select category of genes. Other global signals include cold and heat shock proteins that function to help the cell overcome temperature stress.

Table 8.1 shows examples of global control systems known in Escherichia coli.

Quorum Sensing Quorum sensing (Figure 8.22) allows cells to survey their environment for cells of their own kind and involves the sharing of specific small molecules. Once a sufficient concentration of the signaling molecule is present, specific gene expression is triggered.

Other Mechanisms of Regulation Attenuation

Attenuation is a mechanism whereby gene expression (typically at the level of transcription) is controlled after initiation of RNA synthesis (Figure 8.25).

Attenuation mechanisms involve a coupling of transcription and translation and can therefore occur only in prokaryotes.

Signal Transduction and Two- Component Regulatory Systems

Signal transduction systems transmit environmental signals to the cell.

In prokaryotes, signal transduction typically involves two-component regulatory systems (Figure 8.26), which include a membrane- integrated sensor kinase protein and a cytoplasmic response regulator protein. The activity of the response regulator depends on its state of phosphorylation.

Table 8.3 shows examples of two- component regulatory systems that regulate transcription in Escherichia coli.

Regulation of Chemotaxis Chemotaxis is under complex regulation involving signal transduction in which regulation occurs in the activity of the proteins involved rather than in their synthesis (Figure 8.27).

MCP= methyl accepting chemotaxis proteins

Adaptation by methylation allows the system to reset itself to the continued presence of a signal.

RNA Regulation and Riboswitches RNA regulation is a rapidly expanding area in both prokaryotic and eukaryotic molecular biology.

In Escherichia coli, for example, a number of small RNAs (sRNAs) have been found to regulate various aspects of cell physiology by binding to other RNAs or even to small molecules.

One mechanisms for sRNA activity is found in the signal recognition particle. A second mechanism is the binding of the sRNA to an mRNA by complementary base pairing (Figure 8.28a).

A unique form of small RNAs are the riboswitches. These are mRNAs that contain sequences upstream of the coding sequences that can bind small molecules (Figure 8.28b).

Metabolite binding effects secondary structure of RNA