Laredo J, Wolff V L, Lovett P S
Department of Biological Sciences, University of Maryland Baltimore County, Catonsville 21228.
Gene. 1988 Dec 15;73(1):209-14. doi: 10.1016/0378-1119(88)90327-7.
Gene cat-86 is chloramphenicol (Cm)-inducible and specifies Cm acetyltransferase, CAT-86. The gene was previously cloned from the DNA of a strain of Bacillus pumilus. In the present study we report the construction of a constitutively expressed version of cat-86 that permits high-level expression of the gene on a plasmid in B. subtilis. A method is described that allows very rapid purification of CAT-86 protein to homogeneity. The sequence of 13 N-terminal amino acids of purified CAT-86, as well as the 26.6-kDa size of the subunit protein, agree with predictions made based on the nucleotide sequence of the gene. The Mr of the native enzyme suggests that CAT-86 is a trimer consisting of three identical protein subunits. Our studies demonstrate that cat-86 provides a convenient system for analyzing relationships between a gene and a multimeric enzyme in the B. subtilis background.
基因cat - 86受氯霉素（Cm）诱导，编码氯霉素乙酰转移酶CAT - 86。该基因先前是从短小芽孢杆菌菌株的DNA中克隆出来的。在本研究中，我们报道了构建一个组成型表达的cat - 86版本，它能使该基因在枯草芽孢杆菌的质粒上实现高水平表达。本文描述了一种能非常快速地将CAT - 86蛋白纯化至同质的方法。纯化后的CAT - 86的13个N端氨基酸序列以及亚基蛋白26.6 kDa的大小，与基于该基因核苷酸序列所做的预测相符。天然酶的相对分子质量表明CAT - 86是由三个相同蛋白质亚基组成的三聚体。我们的研究表明，cat - 86为在枯草芽孢杆菌背景下分析基因与多聚体酶之间的关系提供了一个便利的系统。
