Acetyl Coenzyme A Carboxylase
SUBUNIT STRUCTURE OF THE PROTOMERIC FORM OF THE AVIAN LIVER ENZYME
Ras B. Guchhait 1, Eberhard E. Zwergel 1, and M. Daniel Lane 1
From the
1 From the Department of Physiological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
Chicken liver acetyl coenzyme A carboxylase is a polymeric filamentous structure of 4 to 10 million daltons and is composed of linearly aggregated protomers. The protomeric form can be dissociated with sodium dodecyl sulfate and resolved into polypeptide chains of three weight classes (117,000, 129,000, and 139,000 daltons) by polyacrylamide gel electrophoresis in urea-dodecyl sulfate at pH 7.2. The molar stoichiometry based upon relative staining intensity is approximately 2:1:1 for the 117,000-, 129,000-, and 139,000- dalton subunits, respectively, and is consistent with a tetrameric structure of the protomer. Covalently bound biotinyl prosthetic group is associated with only one of the 117,000 molecular weight subunits.
The protomeric form of the carboxylase was cross-linked using the bifunctional reagent, dimethylsuberimidate. Analysis of the cross-linked mixture of polypeptide chains using a low resolution dodecyl sulfate-acrylamide gel electrophoresis system yielded a maximum of four protein bands all of which contain biotin. The molecular weights of these four cross-linked species correspond to those of a monomer (117,000), dimer (234,000), trimer (350,000), and tetramer (470,000). By varying the concentration of dimethylsuberimidate and time of reaction, the distribution of cross-linked polypeptides can be shifted either toward the monomeric or the tetrameric species. These results and the biotin content of the enzyme (1.0 mole per 488,000 g of protein) show the protomeric form to be a tetramer having a molecular weight of 470,000 to 500,000.
Submitted on January 11, 1974
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