Hipps D S, Packman L C, Allen M D, Fuller C, Sakaguchi K, Appella E, Perham R N
Department of Biochemistry, University of Cambridge, U.K.
Biochem J. 1994 Jan 1;297 ( Pt 1)(Pt 1):137-43. doi: 10.1042/bj2970137.
The peripheral subunit-binding domain of the dihydrolipoyl acetyltransferase polypeptide chain of the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus was released by limited proteolysis from a di-domain (lipoyl domain plus binding domain) encoded by a subgene over-expressed in Escherichia coli. The domain was characterized by N-terminal sequence analysis, electrospray m.s. and c.d. spectroscopy. It was found to be identical in all respects to a chemically synthesized peptide of the same sequence. The association of the di-domain and binding domain (both natural and synthetic) with dihydrolipoyl dehydrogenase was analysed in detail and a tight binding was demonstrated. As judged by several different techniques, it was found that only one peripheral subunit-binding domain is bound to one dimer of dihydrolipoyl dehydrogenase, implying that the association is highly anti-cooperative.
嗜热脂肪芽孢杆菌丙酮酸脱氢酶多酶复合体的二氢硫辛酰乙酰转移酶多肽链的外周亚基结合结构域，是通过对在大肠杆菌中过表达的一个亚基因编码的双结构域（硫辛酰结构域加结合结构域）进行有限蛋白酶解而释放出来的。该结构域通过N端序列分析、电喷雾质谱和圆二色光谱进行表征。结果发现它在所有方面都与相同序列的化学合成肽相同。详细分析了双结构域和结合结构域（天然的和合成的）与二氢硫辛酰脱氢酶的结合情况，并证明了紧密结合。通过几种不同技术判断，发现只有一个外周亚基结合结构域与一个二氢硫辛酰脱氢酶二聚体结合，这意味着这种结合具有高度的反协同性。
