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Updated: Mar 19, 2026
Recombinant Protein Expression, Crystallization, and Biophysical Studies of a Bacillus-conserved Nucleotide Pyrophosphorylase, BcMazG
Published on: May 16, 2017
Purification of Pyruvate Dehydrogenase Complex from an Extreme Thermophile, Bacillus caldolyticus, and Its Thermal
Y Hiromasa1, Y Aso1, S Yamashita1,2
1a Laboratory of Protein Chemistry and Engineering, Graduate School of Genetic Resources and Technology, Kyushu University , Higashi-ku, Fukuoka 812 , Japan.
Abstract:
Pyruvate dehydrogenase multienzyme complex was purified from Bacillus caldolyticus. The complex was composed of four polypeptides with molecular masses of 39.8, 41.7, 53.7, and 57.5kDa estimated by SDS-PAGE and they were presumed to be pyruvate decarboxylase (E1, dimeric), lipoate acetyltransferase (E2), and lipoamide dehydrogenase (E3) on the analogy of those from Bacillus stearothermophilus. E1 and E3 were stable at pH 5.7-10.2 and 4.5-11.3, respectively. Halves of E1 and E3 activity were abolished by incubation for 30min at 65°C and 85°C, respectively. Loss of overall activity was principally due to inactivation of E1. Structural changes in the complex incubated at high temperature were studied by fluorescence spectroscopy. The results suggested that the thermal denaturation of the complex proceeded through at least two different steps: inactivations of E1 and E3, and the former process is accompanied by a reduction of the complex size.
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