Cloning of aroG, the gene coding for phospho-2-keto-3-deoxy-heptonate aldolase(phe), in Escherichia coli K-12, and subcloning of the aroG promoter and operator in a promoter-detecting plasmid

Defective transducing phages carrying aroG, the structural gene for phenylalanine (phe)-inhibitable phospho-2-keto-heptonate aldolase (EC 4.1.2.15; previously known as 3-deoxy-D-arabinoheptulosonate-7-phosphate synthetase[phe]), have been isolated, and DNA from two of these phages has been used to construct a restriction map of the region from att lambda to aroG. A 7.6-kb PstI-HindIII fragment from one of these phages was cloned into pBR322 and shown to contain aroG. The location of aroG within the 7.6 kb was established by subcloning and Tn3 transpositional mutagenesis. A fragment carrying the aroG promoter and operator has been cloned into a high copy number promoter-cloning vector (pMC489), and the resulting aroGpo-LacZ' (alpha) fusion subcloned in a low copy number vector. Strains with this fusion on the low copy number vector exhibit negative regulation of beta-galactosidase expression by both phenylalanine and tryptophan and positive regulation by tyrosine in a tyrR+ background.     

UDP-N-acetylglucosamine 1-carboxyvinyl-transferase from Acinetobacter calcoaceticus

Abstract We have analyzed the sequence downstream of rpoN from Zcinetobacter calcoaceticus and identified an open reading frame encoding a protein with high similarity to UDP- N -acetylgucosamine 1-carboxyvinyl-transferase (MurZ). Multicopy plasmids encoding this enzyme conferred phosphomycin resistance to A. calcoaceticus . The polar effect of a rpoN mutation on the phosphomycin resistance level suggests that murZ is, in part, cotranscribed with rpoN . These observations confirm that A. calcoaceticus represents the first exceptin from a conserved genetic context of rpoN observed in several other Gram-negative bacteria.     

蚕豆保卫细胞中ABA响应蛋白基因的转录

植物在不同的逆境条件下可以生成一类受脱落酸（ABA）诱导的蛋白质［脱落酸响应蛋白（ABAresPonsiveProtein，RABpr。tein）］（Bray1993）。RAB蛋白分布在不同的物种之中，许多［如Lea（Lateembryogen－。isabundant）蛋白（Dure1993）］形成于植物胚胎成熟失水过程中，但也有一些是植物受到不同逆境处理后在营养器官内所形成的「如脱水蛋白（Dehrdrin）］（Dure1993）。已发现的70余个RAB蛋白中，有30余个属于脱水蛋白。（Close等1993）RAB蛋白在植物体内的功能目前尚不了解（Bray1993）。由蛋白质的氨基酸组成分析表明，这些…     

Probing the Structural Basis for Enzyme-Substrate Recognition In Cu,Zn Superoxide Dismutase

A full understanding of enzyme-substrate interactions requires a detailed knowledge of their structural basis at atomic resolution. Crystallographic and biochemical data have been analyzed with coupled computational and computer graphic approaches to characterize the molecular basis for recognition of the superoxide anion substrate by Cu. Zn superoxide dismutase (SOD). Detailed analysis of the bovine SOD structure aligned with SOD sequences from 15 species provides new results concerning the significance and molecular basis for sequence conservation. Specific roles have been assigned for all 23 invariant residues and additional residues exhibiting functional equivalence. Sequence invariance is dominated by 15 residues that form the active site stcreochemistry. supporting a primary biological function of superoxide dismutation. Using data from crystallographic structures and site-directed mutants, we are testing the role of individual residues in the active site channel, including (in human SOD) Glu132, Glu133, Lys136, Thr137, and Arg 143. Electrostatic calculations incorporating molecular flexibility suggest that the region of positive electrostatic potential in and over the active site channel above the Cu ion sweeps through space during molecular motion to enhance the facilitated diffusion responsible for the enzyme's rapid catalytic rate.     

Sequence and functional analysis of the cloned Neisseria meningitidis CMP-NeuNAc synthetase

The CMP-N-acetylneuraminic acid (CMP-NeuNAc) synthetase gene of Neisseria meningitidis group B is located on a 2.3-kb EcoRI fragment within the cps gene cluster. Nucleotide sequence determination of the gene encoding the CMP-NeuNAc synthetase revealed a 515-bp open reading frame that can encode a 18.9-kDA protein. A computer data base scan revealed a 59.4% identity to the CMP-NeuNAc synthetase gene of E. coli K1. Enzymatic activity was confirmed in vitro and in vivo. Transformation of the CMP-NeuNAc defective E. coli K1 strain EV5 with the meningococcal CMP-NeuNAc synthetase could complement the defect in E. coli.