Resolution of component proteins in an enzyme complex from Methanosarcina thermophila catalyzing the synthesis or cleavage of acetyl-CoA

Darren R. Abbanat, James G. Ferry

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Abstract

An enzyme complex was isolated from acetate-grown Methanosarcina thermophila that oxidized CO and catalyzed the synthesis or cleavage of acetyl-CoA. The complex consisted of five subunits (α1β1γ1ε1) of 89, 71, 60, 58, and 19 kDa. The complex contained nickel, iron, acid-labile sulfide, and cobalt in a corrinoid cofactor. Two components were resolved by anion-exchange chromatography of the complex in the presence of dodecyltrimethylammonium bromide and Triton X-100: a 200-kDa nickel/iron-sulfur protein with the 89-and 19-kDa (α2εx) subunits and a 100-kDa corrinoid/iron-sulfur protein with the 60- and 58-kDa subunits (γ1δ1). The nickel/iron-sulfur component contained 0.21 Ni, 2.7 Zn, 7.7 Fe, and 13.2 acid-labile sulfide (per α1ε1). The corrinoid/iron-sulfur component contained 0.7 Co, 0.7 factor in {Coα-[α-(5-hydroxybenzimidazolyl)]-Coβ-cyanocobamide}, 3.0 Fe, and 2.9 acid-labile sulfide (γ1δ1). Both components contained iron-sulfur centers. The nickel/iron-sulfur component oxidized CO and reduced methyl viologen or a ferredoxin isolated from M. thermophila. The nickel/iron-sulfur component also oxidized CO and transferred electrons to the corrinoid/iron-sulfur component, reducing the iron-sulfur and Co centers. UV-visible spectroscopy indicated that the reduced corrinoid/iron-sulfur component could be methylated with CH3I. The results suggest that the enzyme complex from M. thermophila contained at least two enzyme components, each with a specific function. The properties of the component enzymes support a mechanism proposed for acetyl-CoA synthesis (or cleavage) by the enzyme complex.

Original languageEnglish (US)
Pages (from-to)3272-3276
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume88
Issue number8
DOIs
StatePublished - 1991

All Science Journal Classification (ASJC) codes

  • General

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