Structural investigation of catalytically modified F120L and F120Y semisynthetic ribonucleases

Protein Sci. 1994 Jan;3(1):39-50. doi: 10.1002/pro.5560030106.


The structures of two catalytically modified semisynthetic RNases obtained by replacing phenylalanine 120 with leucine and tyrosine have been determined and refined at a resolution of 2.0 A (R = 0.161 and 0.184, respectively). These structures have been compared with the refined 1.8-A structure (R = 0.204) of the fully active phenylalanine-containing enzyme (Martin PD, Doscher MS, Edwards BFP, 1987, J Biol Chem 262:15930-15938) and with the catalytically defective D121A (2.0 A, R = 0.172) and D121N (2.0 A, R = 0.186) analogs (deMel VSJ, Martin PD, Doscher MS, Edwards BFP, 1992, J Biol Chem 267:247-256). The movement away from the active site of the loop containing residues 65-72 is seen in all three catalytically defective analogs--F120L, D121A, and D121N--but not in the fully active (or hyperactive) F120Y. The insertion of the phenolic hydroxyl of Tyr 120 into a hydrogen-bonding network involving the hydroxyl group of Ser 123 and a water molecule in F120Y is the likely basis for the hyperactivity toward uridine 2',3'-cyclic phosphate previously found for this analog (Hodges RS, Merrifield RB, 1974, Int J Pept Protein Res 6:397-405) as well as the threefold increase in KM for cytidine 2',3'-cyclic phosphate found for this analog by ourselves.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Aspartic Acid
  • Binding Sites
  • Crystallography, X-Ray
  • Glutamine
  • Histidine
  • Hydrogen Bonding
  • Lysine
  • Molecular Sequence Data
  • Molecular Structure
  • Peptide Fragments / chemistry
  • Protein Conformation
  • Ribonucleases / chemical synthesis
  • Ribonucleases / chemistry*
  • Software
  • Water / chemistry


  • Peptide Fragments
  • Water
  • Glutamine
  • Aspartic Acid
  • Histidine
  • Ribonucleases
  • Lysine