Effect of complexes of ADP and phosphate analogs on the conformation of the Cys707-Cys697 region of myosin subfragment 1

B. C. Phan, Y. M. Peyser, E. Reisler, A. Muhlrad*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

40 Scopus citations

Abstract

Recent crystallographic studies have suggested structural differences between the complexes of S1 · Mg ADP with the phosphate analogs aluminium fluoride (AlF4-), vanadate (VO43-) and beryllium fluoride (BeF(x)) [Fisher, A. J., Smith, C. A., Thoden, J. B., Smith, R., Sutoh, K., Holden, H. M. and Rayment, I. (1995) Biochemistry 34, 8960-8972; Smith, R. and Rayment, I. (1996) Biochemistry 35, 5404-5417]. In this work, chemical modifications, namely labeling of Cys707 (the reactive SH1 thiol) and Cys707-Cys697 (SH1-SH2) cross-linking, were used to compare the S1 · ADP BeF(x), S1 · ADP AlF4- and S1 · ADP · VO43- complexes with specific states of the myosin-ATPase pathway. Modification of Cys707 with the fluorescent monofunctional reagents 7-diethylamino-3-(4'-maleimidylphenyl)-4-methylcoumarin and N-iodoacetyl-N'-(5-sulfo-1-naphtyl)ethylenediamine has shown that the reactivity of the SH1 group depends on the nucleotide bound to S1. The observed rates of Cys707 modification at 20°C lead to the conclusion that S1 · ADP BeF(x) is similar to S1* · ATP, while S1 · ADP · AlF4- and S1 · ADP · VO43- are more similar to S1** · ADP · P(i). The conformations of the analog states were also compared by monitoring the dissociation of the fluorescent nucleotide analog 1-N6'-ethenoadenosine diphosphate (ADP[C2H2]]) from the active site of Cys707-modified (by N-ethylmaleimide) and Cys707-Cys697 cross-linked (by N,N'-p-phenylene dimaleimide) S1 · ADP[C2H2] · AlF4- and S1 · ADP[C2H2] · BeF(x). Our results suggest that the conformations of the S1 · ADP AlF4-, S1 · ADP · VO43- and S1 · ADP · BeF(x) complexes in the Cys707-Cys697 region are distinct from each other, with the former two at least partially resembling the S1** ADP · P(i) state, while the latter is similar to the prehydrolyzed S1* · ATP state.

Original languageEnglish
Pages (from-to)636-642
Number of pages7
JournalEuropean Journal of Biochemistry
Volume243
Issue number3
DOIs
StatePublished - 1997
Externally publishedYes

Keywords

  • Chemical modification
  • Myosin
  • Phosphate analog
  • Subfragment 1
  • Thiol group

Fingerprint

Dive into the research topics of 'Effect of complexes of ADP and phosphate analogs on the conformation of the Cys707-Cys697 region of myosin subfragment 1'. Together they form a unique fingerprint.

Cite this