Glycogen phosphorylase catalytic mechanism

Glycogen phosphorylases belong to the group of vitamin B6 enzymes bearing a catalytic mechanism that involves the participation of the phosphate group of pyridoxal-5 -phosphate (FTP). The proposed mechanism is a concerted one with a front-side attack, as can be seen in Fig. 5 [109]. In the forward direction, e.g.. 

Fig. 5 Catalytic mechanism of glycogen phosphorylases. The reaction scheme accounts for the reversibility of phosphorolysis of oligosaccharides (R) in the presence of orthophosphate (upper half) and primer-dependent synthesis in the presence of glucose-l-phosphate (lower half). PL enzyme-bound pyridoxal BH-f a general base contributed by the enzyme protein. Reprinted with permission from [109]. Copyright 1990 American Chemical Society...

Unlike other pyridoxal phosphate-dependent enzymes, in which it is the carbonyl group that is essential for catalysis, the internal Schiff base between pyridoxal phosphate and lysine in glycogen phosphorylase can be reduced with sodium borohydride without affecting catalytic activity. Thus, while pyridoxal phosphate is essential for phosphorylase activity, it does not act by the same kind of mechanism as in amino acid metabolism. 

The catalytic activity of enzymes is controlled in several ways. Reversible allosteric control is especially important. For example, the first reaction in many biosynthetic pathways is allosterically inhibited by the ultimate product of the pathway. The inhibition of aspartate trans carbamoyl as e by cytidine triphosphate (Section 10.1) is a well-understood example offeedback inhibition. This type of control can be almost instantaneous. Another recurring mechanism is reversible covalent modification. For example, glycogen phosphorylase, the enzyme catalyzing the breakdovm of glycogen, a storage form of sugar, is activated by phosphorylation of a particular serine residue when glucose is scarce (Section 21.2.1). 

Glycogen phosphorylase b is an enzyme that catalyzes the phospho-lytic breakdown of glycogen to glucose-1-phosphate (Equation 18.11 and Figure 18.17). Crystals of the phosphorylase are catalytically active, but the reaction is too fast to study directly by diffraction methods. A gly-cosylic substrate analogue, heptenitol, is slowly converted to heptulose-2-phosphate, presumably by the same mechanism. 

FIGURE 18.17. The use of X-ray diffrciction to follow the course of the reaction of glycogen phosphorylase. Shown is the catalytic mechanism. 

Other proteins involved in glyeogen synthase control are calmodulin, inhibitor-1, and inhibitor-2. Troponin C, when complexed with Ca " ", can activate phosphorylase kinase, but phosphorylase kinase may have little or no role in the inactivation of glycogen synthase in vivo. Inhibitor-1 (M.W. 18,600), when phosphorylated on threonine 35 by cAMP-dependent protein kinase, binds to and inhibits protein phosphatase-1. The concentration of protein phosphatase-1 is lower than that of inhibitors, and its activity is sensitive to the latter, suggesting that inhibitor-1 phosphorylation may be an important regulatory mechanism for this phosphatase. Inhibitor-2 (M.W. 30,500) appears to be an integral part of protein phosphatase-1, forming an inactive 1 1 complex, called Mg +/ATP-dependent phosphatase, with the catalytic subunit of this enzyme. Phosphorylation of inhibitor-2 (also at a threonine residue) activates protein phosphatase-1. This phosphorylation, which requires Mg -ATP, is catalyzed by an activating factor (Fa), GSK-3. 

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