当前位置： SCI文献检索 > BIOCHEMISTRY期刊下所有文献 > Nuclear magnetic resonance and neutron diffraction studies of the complex of ribonuclease A with uridine vanadate, a transition-state analogue.

Nuclear magnetic resonance and neutron diffraction studies of the complex of ribonuclease A with uridine vanadate, a transition-state analogue.

Abstract：

:The complex of ribonuclease A (RNase A) with uridine vanadate (U-V), a transition-state analogue, has been studied with 51V and proton NMR spectroscopy in solution and by neutron diffraction in the crystalline state. Upon the addition of aliquots of U-V at pH 6.6, the C epsilon-H resonances of the two active-site histidine residues 119 and 12 decrease in intensity while four new resonances appear. Above pH 8 and below pH 5, these four resonances decrease in intensity as the complex dissociates. These four resonances are assigned to His-119 and His-12 in protonated and unprotonated forms in the RNase-U-V complex. These resonances do not titrate or change in relative area in the pH range 5-8, indicating a slow protonation process, and the extent of protonation remains constant with ca. 58% of His-12 and ca. 26% of His-119 being protonated. The results of diffraction studies show that both His-12 and His-119 occupy well-defined positions in the RNase-U-V complex and that both are protonated. However, while the classic interpretation of the mechanism of action of RNase based on the proposal of Findlay et al. [Findlay, D., Herries, D. G., Mathias, A. P., Rabin, B. R., & Ross, C. A. (1962) Biochem. J. 85, 152-153] requires both His-12 and His-119 to be in axial positions relative to the pentacoordinate transition state, in the diffraction structure His-12 is found to be in an equatorial position, while Lys-41 is close to an axial position. Hydrogen exchange data show that the mobility and accessibility of amides in the RNase-U-V complex do not significantly differ from what was observed in the native enzyme. The results of both proton NMR in solution and neutron diffraction in the crystal are compared and interpreted in terms of the mechanism of action of RNase.

journal_name

Biochemistry

journal_title

Biochemistry

authors

Borah B,Chen CW,Egan W,Miller M,Wlodawer A,Cohen JS

doi

10.1021/bi00329a038

subject

Has Abstract

pub_date

1985-04-09 00:00:00

pages

2058-67

issue

eissn

0006-2960

issn

1520-4995

journal_volume

pub_type

杂志文章

在线工具

Nuclear magnetic resonance and neutron diffraction studies of the complex of ribonuclease A with uridine vanadate, a transition-state analogue.