University of Illinois Urbana-Champaign

Component C of the Methylcoenzyme M Methylreductase

Hartzell, Patricia Louise

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https://hdl.handle.net/2142/71182

Description

Title
Component C of the Methylcoenzyme M Methylreductase
Author(s)
Hartzell, Patricia Louise
Issue Date
1987
Department of Study
Microbiology
Discipline
Microbiology
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Chemistry, Biochemistry
Abstract
  • Component C, an enzyme in the methylcoenzyme M methylreductase complex of methanogens, has been proposed to contain the catalytic site for methyl group reduction from CH(,3)-S-CH(,2)CH(,2)SO(,3)('-) (methylcoenzyme M). Component C, a hexamer in an (alpha)(,2), (beta)(,2), (gamma)(,2) configuration, contains a chromophore F(,430), coenzyme M and component B (7-mercaptoheptanoylthreonine phosphate). There are two mol of F(,430), a unique nickel tetrapyrrole, per mol of component C. Coenzyme M, the C-1 carrier in the terminal step of methanogenesis, is also present in a ratio of two mol per mol of protein.
  • The role of the nickel tetrapyrrole, F(,430), in catalysis was tested by isolating and purifying F(,430) and each of the subunits of component C. Recovery of maximal enzyme activity required each subunit, F(,430), and coenzyme M. When F(,430) was omitted from the reassociation mixture, the M(,r) 300,000 apoprotein was formed, but was unable to function in the methanogenic assay. Isomers of the nickel tetrapyrrole differed in their ability to reassociate with the enzyme subunits and yielded holoenzyme with altered specific activities. Salt extracted F(,430), a six-coordinate form, gave optimal reassociation and specific activity whereas the four-coordinate heat extracted F(,430) reassociated poorly with the enzyme subunits.
  • The function of the bound C-1 carrier, coenzyme M, in the enzyme was examined with radiolabel tracing during in vitro methanogenesis using a double labeled substrate, C('3)H(,3)-S-CH(,2)CH(,2)('35)SO(,3)('-). The bound form of the coenzyme was found to be HS-CH(,2)CH(,2)SO(,3)('-) and the stoichiometry of 2 mol of HS-CoM per mol of enzyme was confirmed. The incorporation was slow and was not stoichiometric with methane production during catalysis. The incorporation was specific for the thioethanesulfonate moiety of CH(,3)-S-CoM; labeled HS-CoM was not incorporated into the enzyme during methanogenesis.
  • The substrate analogue, bromoethanesulfonate (BES), was found to competitively inhibit methanogenesis from methylcoenzyme M. The target of inhibition was component C.
Graduation Semester
1987
Type of Resource
text
Permalink
http://hdl.handle.net/2142/71182

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Graduate Dissertations and Theses at Illinois PRIMARY
Graduate Theses and Dissertations at Illinois
Dissertations and Theses - Microbiology