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Title:In Vitro Methanogenesis From Formaldehyde. Identification of Three Carbon-1 Intermediates of the Methanogenic Pathway
Author(s):Escalante Semerena, Jorge Carlos
Department / Program:Microbiology
Degree Granting Institution:University of Illinois at Urbana-Champaign
Subject(s):Biology, Microbiology
Abstract:The biosynthesis of CH(,4) from HCHO and by cell-free extracts of Methanobacterium thermoautotrophicum strain (DELTA)H under H(,2) or N(,2) was investigated. Quantitative conversion of HCHO to CH(,4) was achieved under an atmosphere of H(,2) gas. Under a non-reductive atmosphere of N(,2), HCHO underwent disproportionation (oxidation and reduction) events. A CH(,4)/HCHO ratio of 1/2.9 was observed. Cell-free extracts of methanogens able to oxidize formate to CO(,2) and H(,2) (Mc. jannaschii and Mc. voltae) showed a CH(,4)/HCHO ratio of 1/2. Sets of chemical reactions explaining both findings were proposed. In the case of M. thermoautotrophicum, the oxidized C(,1) units were found bond to a carrier later referred to as Formaldehyde Activating Factor (FAF). A spectrophotometric assay for FAF was implemented and used for the purification of the coenzyme. FAF was purified under strictly anoxic conditions to prevent inactivation by O(,2). ('1)H and ('13)C NMR spectrometry and ultraviolet spectroscopy was used to document the binding of C(,1) units of different oxidation states to FAF. Methanogenesis from each derivative of FAF (formyl, methylene, and methyl) was kinetically comparable to CH(,3)-S-CoM. All the derivatives efficiently substituted for CH(,3)-S-CoM in the CH(,3)-S-CoM-stimulated conversion of CO(,2) to CH(,4), the RPG effect. The enzyme responsible for the oxidation of methylene-FAF to formyl-FAF under N(,2) was found to catalyze the reverse reaction under H(,2). This enzyme was proposed to be part of the methanogenic pathway. Initial purification of the enzyme was reported. Component A(,2) of the methylreductase system was involved in the conversion of HCHO to CH(,4) under H(,2). Initial purification of this enzyme was documented. HCHO, HOCH(,2)-S-CoM, thiazolidine, hexamethylene-tetramine, and L-serine required FAF for their conversion to CH(,4). HCHO was found to react chemically with FAF to form methylene-FAF. HOCH(,2)-S-CoM, thiazolidine, and hexamethylenetetramine were found to be in equilbrium with HCHO. This equilibrium was found to be responsible for the methanogenicity of these compounds. In the case of L-serine, the presence of an FAF-dependent transhydroxymethylase in cell-free extracts of M. thermoautotrophicum was proposed. A new model for methanogenesis from H(,2) and CO(,2) proposed.
Issue Date:1983
Description:243 p.
Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1983.
Other Identifier(s):(UMI)AAI8409917
Date Available in IDEALS:2014-12-16
Date Deposited:1983

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