Kozliak E I, Guilloton M B, Gerami-Nejad M, Fuchs J A, Anderson P M
Department of Biochemistry and Molecular Biology, University of Minnesota, Duluth 55812.
J Bacteriol. 1994 Sep;176(18):5711-7. doi: 10.1128/jb.176.18.5711-5717.1994.
Cyanase catalyzes the reaction of cyanate with bicarbonate to give 2CO2. The cynS gene encoding cyanase, together with the cynT gene for carbonic anhydrase, is part of the cyn operon, the expression of which is induced in Escherichia coli by cyanate. The physiological role of carbonic anhydrase is to prevent depletion of cellular bicarbonate during cyanate decomposition due to loss of CO2 (M.B. Guilloton, A.F. Lamblin, E. I. Kozliak, M. Gerami-Nejad, C. Tu, D. Silverman, P.M. Anderson, and J.A. Fuchs, J. Bacteriol. 175:1443-1451, 1993). A delta cynT mutant strain was extremely sensitive to inhibition of growth by cyanate and did not catalyze decomposition of cyanate (even though an active cyanase was expressed) when grown at a low pCO2 (in air) but had a Cyn+ phenotype at a high pCO2. Here the expression of these two enzymes in this unusual system for cyanate degradation was characterized in more detail. Both enzymes were found to be located in the cytosol and to be present at approximately equal levels in the presence of cyanate. A delta cynT mutant strain could be complemented with high levels of expressed human carbonic anhydrase II; however, the mutant defect was not completely abolished, perhaps because the E. coli carbonic anhydrase is significantly less susceptible to inhibition by cyanate than mammalian carbonic anhydrases. The induced E. coli carbonic anhydrase appears to be particularly adapted to its function in cyanate degradation. Active cyanase remained in cells grown in the presence of either low or high pCO2 after the inducer cyanate was depleted; in contrast, carbonic anhydrase protein was degraded very rapidly (minutes) at a high pCO2 but much more slowly (hours) at a low pCO2. A physiological significance of these observations is suggested by the observation that expression of carbonic anhydrase at a high pCO2 decreased the growth rate.
氰酸酶催化氰酸盐与碳酸氢盐反应生成2分子二氧化碳。编码氰酸酶的cynS基因，与编码碳酸酐酶的cynT基因一起，是cyn操纵子的一部分，其表达在大肠杆菌中由氰酸盐诱导。碳酸酐酶的生理作用是防止在氰酸盐分解过程中由于二氧化碳损失导致细胞内碳酸氢盐耗尽（M.B. Guilloton、A.F. Lamblin、E.I. Kozliak、M. Gerami-Nejad、C. Tu、D. Silverman、P.M. Anderson和J.A. Fuchs，《细菌学杂志》175:1443 - 1451，1993年）。一个缺失cynT的突变菌株对氰酸盐抑制生长极其敏感，并且在低pCO₂（空气中）生长时不催化氰酸盐分解（即使表达了活性氰酸酶），但在高pCO₂时具有Cyn⁺表型。在此，对这两种酶在这种不寻常的氰酸盐降解系统中的表达进行了更详细的表征。发现这两种酶都位于细胞质中，并且在存在氰酸盐的情况下以大致相等的水平存在。一个缺失cynT的突变菌株可以用高水平表达的人碳酸酐酶II进行互补；然而，突变缺陷并未完全消除，可能是因为大肠杆菌碳酸酐酶比哺乳动物碳酸酐酶对氰酸盐抑制的敏感性显著更低。诱导的大肠杆菌碳酸酐酶似乎特别适应其在氰酸盐降解中的功能。在诱导剂氰酸盐耗尽后，活性氰酸酶仍保留在低pCO₂或高pCO₂条件下生长的细胞中；相反，碳酸酐酶蛋白在高pCO₂时降解非常迅速（几分钟），但在低pCO₂时降解慢得多（数小时）。这些观察结果的生理意义通过在高pCO₂时碳酸酐酶表达降低生长速率这一观察结果得以体现。
