Sung Y C, Fuchs J A
Department of Biochemistry, University of Minnesota, St. Paul 55108.
J Biol Chem. 1988 Oct 15;263(29):14769-75.
Escherichia coli can overcome the toxicity of environmental cyanate by hydrolysis of cyanate to ammonia and bicarbonate. This reaction is catalyzed by the enzyme cyanase, encoded by the cynS gene. The nucleotide sequence of cynS has been reported (Sung, Y.-c., Anderson, P. M., and Fuchs, J. A. (1987) J. Bacteriol. 169, 5224-5230). The nucleotide sequence of the complete cyn operon has now been determined. The cyn operon is approximately 2600 base pairs and includes cynT, cynS, and cynX, which encode cyanate permease, cyanase, and a protein of unknown function, respectively. Two cyanate-inducible transcripts of 1500 and 2500 nucleotides, respectively, were detected by Northern blot analysis. S1 nuclease mapping experiments indicated that two different cyn mRNAs have a common 5'-end and two different 3'-ends. One 3'-end was located within the coding region of cynX, whereas the other 3'-end includes the entire DNA sequence of cynX. The longer transcript contained 98 nucleotides complementary to lac mRNA produced by the predominant lac transcription termination sequence. Termination vectors were used to show that both 3'-ends were generated by sequences that caused transcriptional termination in vivo. Expression vectors were used to demonstrate that a protein corresponding to the expected size was synthesized from the DNA fragment containing the open reading frame designated cynX. The predicted amino acid sequence of cynX indicates that it is a very hydrophobic protein. The level of cynX expression was significantly less than that of cynT or cynS expression.
大肠杆菌可通过将氰酸盐水解为氨和碳酸氢盐来克服环境中氰酸盐的毒性。该反应由cynS基因编码的氰酸酶催化。cynS的核苷酸序列已有报道（Sung, Y.-c., Anderson, P. M., and Fuchs, J. A. (1987) J. Bacteriol. 169, 5224 - 5230）。现在已确定了完整的cyn操纵子的核苷酸序列。cyn操纵子约2600个碱基对，包括cynT、cynS和cynX，它们分别编码氰酸盐通透酶、氰酸酶和一种功能未知的蛋白质。通过Northern印迹分析检测到分别为1500和2500个核苷酸的两种氰酸盐诱导转录本。S1核酸酶图谱实验表明，两种不同的cyn mRNA有一个共同的5′末端和两个不同的3′末端。一个3′末端位于cynX的编码区内，而另一个3′末端包括cynX的整个DNA序列。较长的转录本包含98个与由主要的lac转录终止序列产生的lac mRNA互补的核苷酸。使用终止载体表明，两个3′末端均由体内导致转录终止的序列产生。使用表达载体证明，从包含指定为cynX的开放阅读框的DNA片段合成了与预期大小相对应的蛋白质。cynX的预测氨基酸序列表明它是一种非常疏水的蛋白质。cynX的表达水平明显低于cynT或cynS的表达水平。
