Stauffer L T, Ghrist A, Stauffer G V
Department of Microbiology, University of Iowa, Iowa City 52242.
DNA Seq. 1993;3(6):339-46. doi: 10.3109/10425179309020835.
Plasmid pGS146 carries the Escherichia coli gcv system on a 7.12 kb SalI-BamHI DNA insert fragment. The DNA sequence of a gene which presumably encodes the T-protein of the glycine cleavage (GCV) enzyme complex was determined. The gene, designated gcvT, encodes a polypeptide of 364 amino acids with a calculated molecular weight of 40,146 daltons. In a minicell system, the SalI-BamHI fragment directs the synthesis of three polypeptides with Mr values of about 93,300, 43,300 and 17,400 daltons. When gcvT was inactivated by insertion of a translation terminator sequence, the Mr 43,300 dalton polypeptide was not observed. The deduced amino acid sequence of the E. coli T-protein was compared with the sequence of the T-protein from bovine liver. 190 of 364 amino acid residues are identical or chemically similar between the two proteins. An S1 nuclease mapping experiment located the transcription start point for gcvT. Single basepair changes were made in the promoter -10 and -35 sequences. These mutations significantly reduced expression from a gcvT-lacZ gene fusion. The gcvT gene is transcribed and translated in the same direction as the gcvH gene.
质粒pGS146在一个7.12 kb的SalI - BamHI DNA插入片段上携带大肠杆菌的gcv系统。测定了一个可能编码甘氨酸裂解（GCV）酶复合物T蛋白的基因的DNA序列。该基因命名为gcvT，编码一个由364个氨基酸组成的多肽，计算分子量为40,146道尔顿。在一个小细胞系统中，SalI - BamHI片段指导合成三种分子量约为93,300、43,300和17,400道尔顿的多肽。当通过插入翻译终止序列使gcvT失活时，未观察到分子量为43,300道尔顿的多肽。将大肠杆菌T蛋白的推导氨基酸序列与牛肝T蛋白的序列进行了比较。两种蛋白质的364个氨基酸残基中有190个相同或化学性质相似。一项S1核酸酶图谱实验确定了gcvT的转录起始点。在启动子的 - 10和 - 35序列中进行了单碱基对改变。这些突变显著降低了gcvT - lacZ基因融合体的表达。gcvT基因与gcvH基因的转录和翻译方向相同。
