The Structure and Function of Ribonuclease T1

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Abstract
1. In order to obtain some information on the role of the single arginine residue at position 77 in the enzymatic activity of ribonuclease T1 [EC 2.7.7.26], the enzyme was allowed to react at pH 8.0 and 25°C with phenylglyoxal and glyoxal, reagents known to modify arginine residues in proteins fairly specifically. When the enzyme (0.16% solution) was treated with a 760-fold molar excess of phenylglyoxal, activity toward both RNA and 2′, 3′-cyclic guanylic acid was lost in parallel. Under this condition the half life was about 4hr. A similar inactivation occurred when the enzyme was treated with glyoxal. In this case, however, the activity toward RNA was lost somewhat faster than that toward 2′, 3′-cyclic guanylate. Inactivation was slower in the presence of 3′-guanylic acid. 2. Amino acid analyses of acid hydrolysates of the inactivated proteins showed that the arginine residue and one alanine residue (presumably the amino-terminal residue) had been lost and that the loss of activity almost paralleled the loss of the arginine residue. The reactivity of the active site glutamic acid-58 toward iodoacetate was also decreased almost in parallel with the loss of the arginine residue by reaction with phenylglyoxal. 3. These lines of evidence seem to indicate that the single arginine residue in ribonuclease T1 is present at or near the active center of the enzyme and may be part of the binding site or involved in building the architecture of the active center of the enzyme.

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