Phosphorylase blocks

Showdomycin. Showdomycin (2-p-D-ribofuranosyhnaleimide) (7) is a maleimide C-nucleoside antibiotic synthesi2ed by S. showdoensis-, isoshowdomycin (8) and maleimycin (9) have also been isolated (1—6). Showdomycin is not phosphorylated by nucleoside kinase and is not a substrate for nucleoside phosphorylase. Once (7) enters the cell, it blocks the uptake of glucose and other nutrients. 

For the hydrosilylation reaction various rhodium, platinum, and cobalt catalysts were employed. For the further chain extension the OH-functionalities were deprotected by KCN in methanol. The final step involved the enzymatic polymerization from the maltoheptaose-modified polystyrene using a-D-glucose-l-phosphalc dipotassium salt dihydrate in a citrate buffer (pH = 6.2) and potato phosphorylase (Scheme 59). The characterization of the block copolymers was problematic in the case of high amylose contents, due to the insolubility of the copolymers in THF. 

Figure 2.5. Diagramatic representation of the goal of rational drug design (a) illustrates the normal catalytic activity exhibited by purine nucleoside phosphorylase (PNP) (b) represents an effective inhibitor of PNP, which hts well into the active site thereby blocking its normal enzymatic activity...

The polypeptide of type-L isozyme is composed of 916 amino acid residues. There are two covalently modified amino adds about one-fourth of the amino-terminal threonines are blocked by an acetyl group, and a lysyl residue (Lys762) is linked to the cofactor pyridoxal-P. The partial amino-terminal acetylation appeared to be a natural feature of type-L isozyme, and both the blocked and unblocked forms of type-L phosphorylase may exist in potato tubers. 

Figure 3. Contour plot of an LC/MS of a chymotryptic digest of modifled phosphorylase b, printed in black and white mode. Circles indicate the positions of peaks from a contour plot of the le-acetylated digest for comparison. The two significant shared peaks (indicated by arrows) are the N-terminal blocked peptide (at m/z=514 and 13 min elution time) and the leucine enkephalin internal standard (at m/z=SS6 and 21 min elution time).

Comparison of the structures of phosphorylase a and phosphorylase b reveals that subtle structural changes at the subunit interfaces are transmitted to the active sites (see Figure 21.9). The transition from the T state (represented by phosphorylase b) to the R state (represented by phosphorylase a) entails a 10-degree rotation around the twofold axis of the dimer. Most importantly, this transition is associated with structural changes in a helices that move a loop out of the active site of each subunit. Thus, the T state is less active because the catalytic site is partly blocked. In the R state, the catalytic site is more accessible and a binding site for orthophosphate is well organized. 

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