Vsj. Demel et al., STRUCTURAL INVESTIGATION OF CATALYTICALLY MODIFIED F120L AND F120Y SEMISYNTHETIC RIBONUCLEASES, Protein science, 3(1), 1994, pp. 39-50
The structures of two catalytically modified semisynthetic RNases obta
ined by replacing phenylalanine 120 with leucine and tyrosine have bee
n determined and refined at a resolution of 2.0 Angstrom (R = 0.161 an
d 0.184, respectively). These structures have been compared with the r
efined 1.8-Angstrom structure (R = 0.204) of the fully active phenylal
anine-containing enzyme (Martin PD, Doscher MS, Edwards BFP, 1987, J B
iol Chem 262:15930-15938) and with the catalytically defective D121A (
2.0 Angstrom, R = 0.172) and D121N (2.0 Angstrom, R = 0.186) analogs (
deMel VSJ, Martin PD, Doscher MS, Edwards BFP, 1992, J Biol Chem 267:2
47-256). The movement away from the active site of the loop containing
residues 65-72 is seen in ah 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 hydro
gen-bonding network involving the hydroxyl group of Ser 123 and a wate
r molecule in F120Y is the likely basis for the hyperactivity toward u
ridine 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 K-M for cytidine 2',3'-cyclic phosphate fou
nd for this analog by ourselves.