regulatory strategies enzymes and hemoglobin. regulatory strategies what are the four ways in which...
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Regulatory Strategies
What are the four ways in which enzymes are regulated?– allosteric regulation– enzymes exist in multiple forms (isoenzymes)– covalent modification– proteolytic cleavage
Allosteric Regulation
What are some of the characteristics of this form of regulation?– Activity influenced by non-covalent binding of
metabolite called a modulator• May be inhibitory or stimulatory
• May have one (monovalent) or several (polyvalent) modulators
• Binding induces shape change in enzyme
• Enzymes are large; two or more subunits
• Exhibit homotropic or heterotropic control
Allosteric Regulation
– enzyme may be part of a sequence in which the end product inhibits allosteric enzyme
– enzyme does not show Michaelis-Menten kinetics
Aspartate Transcarbamoylase
CTP is a negative modulator and ATP is a positive modulator– feedback inhibition
Aspartate Transcarbamoylase
How do we know that the catalytic and regulatory sites on this enzyme are distinct?– treat with
p-hydroxymercuribenzoate
Aspartate Transcarbamoylase
Catalytic subunit consists of 3 chains (c3) and regulatory subunit consists of 2 chains (r2)
Aspartate Transcarbamoylase
What is the difference between the T state and the R state of the enzyme?– T = tense; lower affinity for substrate– R = relaxed; higher affinity for substrate
Binding of substrate or substrate analog converts enzyme from T to R state– positive cooperativity– homotropic control
Aspartate Transcarbamoylase
How does CTP act as a negative modulator?
Why is this an example of heterotropic modulation?
Aspartate Transcarbamoylase
How does ATP act as a positive modulator?
Does this represent homo or heterotropic modulation?
Aspartate Transcarbamoylase
Why is the mechanism just described called the concerted model?– All subunits must be
in same conformation, T or R
Allosteric Regulation
How does the sequential model differ from the concerted model?– Subunits may undergo individual sequential changes
in conformation
– Subunits can interact even in different conformations
– Change induced by binding of substrate to one subunits can increase or decrease substrate binding to other subunits
• positive or negative homotropic effects
• finer tuning
Allostery and Hemoglobin
Binding of O2 to hemoglobin represented by a sigmoidal curve similar to allosteric enzymes
Cooperativity promotes efficient O2 delivery
Allostery and Hemoglobin
How does 2,3 bisphosphoglycerate affect O2 affinity of hemoglobin?
How is this effect brought about?
Allostery and Hemoglobin
How is fetal hemoglobin different from maternal hemoglobin?
How can this be explained?
Isozymes
What are isozymes or isoenzymes?– enzymes that have differences in amino acid
sequence but catalyze the same reaction• Example – lactate dehydrogenase
• Two different chains, M and H, exist
• Enzyme consists of 4 subunits– H4, H3M, H2M2, HM3, M4
• Each form has different Km and Vmax
Covalent Modification
Enzymes exist in active and inactive forms– Interconvertable by covalent modification
• Catalyzed by other enzymes
• Most modifications are reversible
Covalent Modification
What are the most common forms of covalent modification?– Phosphorylation and dephosphorylation
Which enzymes catalyze phosphorylation?– protein kinases
Which enzymes catalyze dephosphorylation?– protein phosphatases
Covalent Modification
Why is phosphorylation an effective way to regulate proteins?– Phosphate group adds negative charges– Phosphate group can form hydrogen bonds– Free energy of phosphorylation is large– Can occur rapidly or slowly as needed– Can achieve amplification– Linked to energy status of cell
Covalent Modification
Example – glycogen phosphorylase– Two forms a and b
• A = active; 4 subunits each with a serine residue phosphorylated at OH group
• B = inactive; removal of PO4 groups causes protein to separate into two half molecules
Covalent Modification
What is the connection between c-AMP and protein kinases?– PKA activated by c-AMP in cells
Zymogens
What are zymogens?– inactive precursors of enzymes
How are they activated?– proteolytic cleavage
Zymogens
What are some other examples?– Blood clotting– Insulin– Conversion of procollagenase to collaginase in
metamorphosis– Conversion of procaspases to caspases in
apoptosis
Zymogens
Why do proteolytic enzymes have specific inhibitors?– To prevent premature activation of the enzyme– Activation of trypsin in pancreas could destroy
pancreatic tissue