The reactivity of arginine residues interacting with glucose 1-phosphate in glycogen phosphorylase. A comparison between pyridoxal-reconstituted phosphorylase and the native enzyme.

Modification of pyridoxal-reconstituted phosphorylase b with two arginine-directed reagents, butanedione and [14C]phenylglyoxal, has been investigated and compared with the results obtained on the active and inactive conformations of the native enzyme; the reactivity of the various arginine residues has been directly described using autoradiography of chymotryptic maps derived from [14C]phenylglyoxal-labelled phosphorylase. In the native enzyme this method demonstrates that the same arginine residue (568) is reactive on both activated phosphorylase a and b, non-reactive on inactive forms of phosphorylase and protected by glucose 1-phosphate. Another residue is reactive, but its reactivity does not drastically depend upon phosphorylase conformation; it interacts with glucose 1-phosphate. In the pyridoxal-reconstituted phosphorylase, the residue Arg-568 is reactive. This reactivity does not correlated in a simple manner with the ionisation state of the coenzyme, since it is high when this group is either absent or in a dianionic form, and low when it is monoanionic. The reactivity of Arg-568 rather correlates with the quaternary structure of the enzyme. The protection offered by glucose 1-phosphate, pyrophosphate and phosphite on this pyridoxal-reconstituted phosphorylase also provides information about the relative disposition of the substrate, the coenzyme and this particular arginine residue.