6RSA

NUCLEAR MAGNETIC RESONANCE AND NEUTRON DIFFRACTION STUDIES OF THE COMPLEX OF RIBONUCLEASE*A WITH URIDINE VANADATE, A TRANSITION-STATE ANALOGUE

6RSA の概要

• エントリーDOI: 10.2210/pdb6rsa/pdb
• 分子名称: RIBONUCLEASE A, URIDINE-2',3'-VANADATE (3 entities in total)
• 機能のキーワード: hydrolase
• 由来する生物種: Bos taurus (cattle)
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 14051.47

構造登録者

Wlodawer, A. (登録日: 1986-02-25, 公開日: 1986-05-07, 最終更新日: 2024-10-30)

主引用文献

Borah, B.,Chen, C.W.,Egan, W.,Miller, M.,Wlodawer, A.,Cohen, J.S.
Nuclear magnetic resonance and neutron diffraction studies of the complex of ribonuclease A with uridine vanadate, a transition-state analogue.
Biochemistry, 24:2058-2067, 1985

PubMed 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.

PubMed: 4016100
DOI: 10.1021/bi00329a038

実験手法

NEUTRON DIFFRACTION (2 Å)
SOLUTION NMR