Evolutionary Relationships among Bacterial Carbamoyltransferases

Abstract

An immunological approach was used for the study of ornithine carbamoyltransferase (OTCase) evolution in bacteria. Antisera were prepared against the anabolic and catabolic OTCases of Pseudomonas aeruginosa and Aeromonas formicans as well as against OTCase and putrescine carbamoyltransferases from Streptococcus faecalis; these antisera were then tested against the unpurified OTCases, either anabolic or catabolic, of 34 bacterial strains. Extensive crossreactions were observed between the antisera to catabolic OTCases from P . aeruginosa, A . formicans and S . faecalis and the catabolic enzymes from other species or genera. These antisera cross-reacted also with the anabolic OTCases of strains of the Enterobacteriaceae but not with the anabolic OTCases of the same species or of other species or genera. The cross-reaction measured between the antisera against P. aeruginosa anabolic OTCase and the anabolic OTCases of other Pseudomonas were largely in agreement with the phylogenic subdivision of Pseudomonas proposed by N. J. Palleroni. The correlation was also significantly higher with the anabolic enzyme of an archaeobacterium, Methanobacterium thermoaceticum, than with the catabolic or anabolic OTCases from other genera in the eubacterial line. The antiserum raised against A . formicans anabolic OTCase was quite specific for its antigen and appeared to be raised against the heaviest of the various oligomeric structures of the enzyme. INTRODUCTIONAnabolic ornithine carbamoyltransferase (OTCase; EC 2.1 .3.3) functions in the biosynthesis of arginine and catalyses the formation of citrulline and phosphate from ornithine and carbamoylphosphate. The anabolic OTCase from a variety of organisms occurs as a trimeric molecule with a subunit molecular mass ranging from 35 to 40 kDa. This trimeric structure also exists in other enzymes involved in carbamoylphosphate metabolism, namely in aspartate carbamoyltransferase (ATCase; EC 2.1 .3.2) and in putrescine carbamoyltransferase (PTCase; EC 2.1.3.6) (Vickers, 1981;Cunin et al., 1986).Catabolic OTCase is part of the catabolic arginine deiminase pathway and catalyses the formation of carbamoylphosphate and ornithine from citrulline and phosphate. Catabolic OTCases are usually large proteins composed of three, six, eight, nine or more identical subunits (Legrain et al., 1977;Marshall & Cohen, 1972;Baur et al., 1987).Arginine prototrophic organisms that possess the arginine deiminase pathway have two OTCases, one anabolic and the other catabolic. In Pseudomonas putida and Pseudomonas aeruginosa, both enzymes function unidirectionally. The Pseudomonas catabolic enzymes display intense allosteric behaviour which probably results from their large oligomeric structure, , 1987). In this context it is not clear whether trimeric and multimeric OTCases are similar proteins resulting from convergence, or homologous proteins resulting from the differentiation and specialization of an ancestral carbamoyltransferase.To answer this question we have determined the amino acid sequence of the t...

Acetylornithine Transcarbamylase: a Novel Enzyme in Arginine Biosynthesis

“…One of the essential enzymes in the pathway, ornithine transcarbamylase (OTCase) (EC 2.1.3.3), catalyzes the formation of citrulline from ornithine and carbamyl phosphate. Protein sequences within the OTCase family show strong conservation across diverse phylogenetic domains ranging from archaea to mammals (25,26). This suggests that the role of OTCase in arginine biosynthesis was established at a very early stage of evolution, an assertion that is further supported by phylogenetic comparison of the OTCase sequences to the paralogous aspartate transcarbamylases (ATCases) involved in pyrimidine synthesis.…”

The Gene Cluster for Agmatine Catabolism ofEnterococcus faecalis: Study of Recombinant Putrescine Transcarbamylase and Agmatine Deiminase and a Snapshot of Agmatine Deiminase Catalyzing Its Reaction

“…Thus, this gene cluster would contain the genes for all the catalysts required for operation of the AgDI pathway, and indeed, a polar disruption of the first gene in this cluster of S. mutans strongly decreased the AgDI activity measured in permeabilized cells, as expected for the AgDI operon (17). Further, the amino acid sequence predicted to be encoded by ef0732 coincides with the N-terminal sequence reported long ago for E. faecalis PTC (39,54).…”

Catabolic ornithine transcarbamylase of Halobacterium halobium (salinarium): purification, characterization, sequence determination, and evolution

“…Therefore, within the OTCase protein family the anabolic OTCase of P. aeruginosa and the cOTCase of H. salinarium deviate from the evolution of organisms. It has been shown before that the similarities between OTCases is not straightforward but, e.g., on the basis of immunological methods, the catabolic enzymes from P. aeruginosa and Aeromonas formicans are more related to the anabolic enzyme of E. coli than to the anabolic enzymes from the respective organisms (14,52).…”