Hong J, Salo W L, Lusty C J, Anderson P M
Department of Biochemistry and Molecular Biology, University of Minnesota, Duluth 55812.
J Mol Biol. 1994 Oct 14;243(1):131-40. doi: 10.1006/jmbi.1994.1638.
The amino acid sequence of carbamyl phosphate synthetase (CPS) III from liver of spiny dogfish shark Squalus acanthias was deduced from the nucleotide sequence of its cDNA. Alignment of the derived amino acid sequence of CPS III with sequences of rat and frog CPS I and hamster CPS II reveals a high degree of amino acid identity, indicating that CPS III shares the same common ancestral genes as CPSs I and II. All of the CPSs examined show a high conservation of sequences in the adenine nucleotide binding domains and in residues that have been implicated in catalysis. The active-site cysteine residue required for glutamine-dependent activity by CPS II is preserved in the sequence of CPS III. Nevertheless, analysis of the protein sequences indicates that CPS III is more closely related to CPS I than to CPS II. The structure of CPS III, which is composed of a single polypeptide, is consistent with the view that CPS III evolved by fusion of separate genes coding for the glutaminase and synthetase domains of the enzyme and, like other CPSs, the synthetase domain evolved by duplication and fusion of an ancestral kinase gene. These results, together with the recent finding that frog CPS I retains the active site cysteine residue in the glutaminase domain required for glutamine-dependent activity, indicate that other amino acid substitutions critical for glutamine-dependent activity preceded loss of this catalytic cysteine residue. The results described here together with earlier biochemical evidence support the view that acetylglutamate and glutamine-dependent CPS III found in invertebrates and fish species represents an intermediate in the evolution of ancestral glutamine-dependent CPS II toward the acetylglutamate and ammonia-dependent CPS I of ureotelic terrestrial vertebrates.
通过对白斑角鲨肝脏中氨甲酰磷酸合成酶（CPS）III的cDNA核苷酸序列进行推导，得出了其氨基酸序列。将推导得到的CPS III氨基酸序列与大鼠和青蛙的CPS I以及仓鼠的CPS II序列进行比对，发现氨基酸具有高度同一性，这表明CPS III与CPS I和CPS II拥有相同的共同祖先基因。所有检测的CPS在腺嘌呤核苷酸结合结构域以及与催化作用相关的残基中都表现出高度的序列保守性。CPS II中谷氨酰胺依赖性活性所需的活性位点半胱氨酸残基在CPS III序列中得以保留。然而，蛋白质序列分析表明，CPS III与CPS I的关系比与CPS II更为密切。CPS III由单一多肽组成，其结构与以下观点一致：CPS III是通过融合编码该酶谷氨酰胺酶和合成酶结构域的独立基因进化而来，并且与其他CPS一样，合成酶结构域是通过祖先激酶基因的复制和融合进化而来。这些结果，连同最近发现青蛙CPS I在谷氨酰胺依赖性活性所需的谷氨酰胺酶结构域中保留了活性位点半胱氨酸残基，表明在该催化半胱氨酸残基丢失之前，还有其他对谷氨酰胺依赖性活性至关重要的氨基酸取代。这里描述的结果与早期的生化证据共同支持了这样一种观点：在无脊椎动物和鱼类中发现的乙酰谷氨酸和谷氨酰胺依赖性CPS III代表了祖先谷氨酰胺依赖性CPS II向排尿素陆生脊椎动物的乙酰谷氨酸和氨依赖性CPS I进化过程中的一个中间阶段。
