Tryptophanase from Bacillus alvei
I. SUBUNIT STRUCTURE
Sallie O'Neil Hoch 1 and R. D. DeMoss 1
From the
1 From the Department of Microbiology, University of Illinois, Urbana, Illinois 61801
Ultracentrifugal analyses of native holotryptophanase of Bacillus alvei demonstrate that it has a molecular weight of 208,000 and a sedimentation coefficient of 9.7 S at infinite dilution. The tryptophanase molecule dissociates in the presence of 5 m guanidine hydrochloride or 8 m urea, plus thiols, to produce four monomers homogeneous by the criterion of weight. The subunit molecular weight was also estimated by sodium dodecyl sulfate polyacrylamide electrophoresis following performic acid oxidation of the enzyme, borohydride reduction after incubation with sodium dodecyl sulfate, and treatment with 1 m hydroxylamine at pH 10. The apparent monomer molecular weight was calculated to be 50,500 ± 1800. Ultracentrifugal studies with the apoenzyme indicate that pyridoxal 5'-phosphate stabilizes the tetrameric form of the enzyme. Reversible dissociation of the apoenzyme is effected by lowering the protein concentration below 3 mg per ml or by raising the pH above 6. Tryptic peptide maps support the hypothesis that there are four identical monomer units, which may in turn contain a high degree of internal sequence homology.
Submitted on August 9, 1971
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