Decomposition of carbon monoxide dehydrogenase into alpha metallosubunits and a catalytically-active form consisting primarily of beta metallosubunits.
Carbon monoxide dehydrogenase from Clostridium thermoaceticum, with an alpha 3 beta 3 quaternary structure, was incubated with dithiothreitol and 740-78,000 equiv/alpha 3 beta 3 of sodium dodecyl sulfate (SDS), followed by electrophoresis under anaerobic native conditions. Three catalytically-active bands and four inactive bands were obtained, in proportions dependent on the amount of SDS added. The catalytically-active bands, called SM-CODH, NM-CODH, and FM-CODH, migrated slower than, similar to, and faster than native enzyme, respectively. Two-dimensional electrophoresis revealed that SM-CODH and NM-CODH contained approximately equal proportions of the alpha and beta subunits, while the beta/alpha ratio for FM-CODH was about 2.7. The four inactive bands were identified as the beta subunit, two forms of the alpha subunit (called alpha and alpha'), and a form that migrated similarly to native enzyme. Overloaded gels revealed that alpha and each active band had brown color, indicating intact Fe-S clusters in these species. Higher concentrations of SDS (> 1600 equiv/alpha 3 beta 3) and/or incubation at temperatures > 15 degrees C yielded more alpha and beta subunits at the expense of the catalytically-active bands. When incubated at 70 degrees C in 78,000 equiv/alpha 3 beta 3 of SDS, alpha quantitatively converted to alpha', suggesting that alpha' is a denatured form. FM-CODH appears to be an intermediate in the decomposition of CODH by SDS and to have either an alpha 1 beta 3 or alpha 1 beta 2 quaternary structure.(ABSTRACT TRUNCATED AT 250 WORDS)
Xia
J
J
Department of Chemistry, Texas A & M University, College Station 77843, USA.
Lindahl
P A
PA
D000445:Aldehyde Oxidoreductases / Q000737:chemistry*; D003013:Clostridium / Q000201:enzymology; D007506:Iron-Sulfur Proteins / Q000737:chemistry; D009097:Multienzyme Complexes; D011487:Protein Conformation; D011489:Protein Denaturation; D012967:Sodium Dodecyl Sulfate
D007506:Iron-Sulfur Proteins; D009097:Multienzyme Complexes; D012967:Sodium Dodecyl Sulfate; D000445:Aldehyde Oxidoreductases; C019823:carbon monoxide dehydrogenase