Aspartate ammonia-lyase

About: Aspartate ammonia-lyase is a research topic. Over the lifetime, 70 publications have been published within this topic receiving 1568 citations. The topic is also known as: ApsA & IPR004708.

Amino acid-dependent growth of Campylobacter jejuni: key roles for aspartase (AspA) under microaerobic and oxygen-limited conditions and identification of AspB (Cj0762), essential for growth on glutamate

Abstract: Amino acids are key carbon and energy sources for the asaccharolytic food-borne human pathogen Campylobacter jejuni. During microaerobic growth in amino acid rich complex media, aspartate, glutamate, proline and serine are the only amino acids significantly utilized by strain NCTC 11168. The catabolism of aspartate and glutamate was investigated. An aspartase (aspA) mutant (unable to utilize any amino acid except serine) and a Cj0762c (aspB) mutant lacking aspartate:glutamate aminotransferase (unable to utilize glutamate), were severely growth impaired in complex media, and an aspA sdaA mutant (also lacking serine dehydratase) failed to grow in complex media unless supplemented with pyruvate and fumarate. Aspartase was shown by activity and proteomic analyses to be upregulated by oxygen limitation, and aspartate enhanced oxygen-limited growth of C. jejuni in an aspA-dependent manner. Stoichiometric aspartate uptake and succinate excretion involving the redundant DcuA and DcuB transporters indicated that in addition to a catabolic role, AspA can provide fumarate for respiration. Significantly, an aspA mutant of C. jejuni 81-176 was impaired in its ability to persist in the intestines of outbred chickens relative to the parent strain. Together, our data highlight the dual function of aspartase in C. jejuni and suggest a role during growth in the avian gut.

Influence of increased aspartate availability on lysine formation by a recombinant strain of Corynebacterium glutamicum and utilization of fumarate.

Abstract: Aspartate availability was increased in Corynebacterium glutamicum strains to assess its influence on lysine production. Upon addition of fumarate to a strain with a feedback-resistant aspartate kinase, the lysine yield increased from 20 to 30 mM. This increase was accompanied by the excretion of malate and succinate. In this strain, fumaric acid was converted to aspartate by fumarate hydratase, malate dehydrogenase, and aspartate amino transferase activity. To achieve the direct conversion of fumarate to aspartate, shuttle vectors containing the aspA+ (aspartase) gene of Escherichia coli were constructed. These constructions were introduced into C. glutamicum, which was originally devoid of the enzyme aspartase. This resulted in an aspartase activity of 0.3 U/mg (70% of the aspartase activity in E. coli) with plasmid pZ1-9 and an activity of up to 1.05 U/mg with plasmid pCE1 delta. In aspA+-expressing strains, lysine excretion was further increased by 20%. Additionally, in strains harboring pCE1 delta, up to 27 mM aspartate was excreted. This indicates that undetermined limitations in the sequence of reactions from aspartate to lysine exist in C. glutamicum.

The structure of L-aspartate ammonia lyase from escherichia coli

Abstract: The X-ray crystal structure of l-aspartate ammonia-lyase has been determined to 2.8 A resolution. The enzyme contains three domains, and each domain is composed almost completely of alpha helices. The central domain is composed of five long helices. In the tetramer, these five helices form a 20-helix cluster. Such clusters have also been seen in delta-crystallin and in fumarase. The active site of aspartase has been located in a region that contains side chains from three different subunits. The structure of the apoenzyme has made it possible to identify some of the residues that are involved in binding the substrate. These residues have been examined by site-directed mutagenesis, and their putative roles have been assigned [Jayasekera, M. M. K., Shi, W., Farber, G. K., & Viola, R. E. (1997) Biochemistry 36, 9145-9150].

L-aspartase: new tricks from an old enzyme.

Abstract: The enzyme L-aspartate ammonia-lyase (aspartase) catalyzes the reversible deamination of the amino acid L-aspartic acid, using a carbanion mechanism to produce fumaric acid and ammonium ion. Aspartase is among the most specific enzymes known with extensive studies failing, until recently, to identify any alternative amino acid substrates that can replace L-aspartic acid. Aspartases from different organisms show high sequence homology, and this homology extends to functionally related enzymes such as the class II fumarases, the argininosuccinate and adenylosuccinate lyases. The high-resolution structure of aspartase reveals a monomer that is composed of three domains oriented in an elongated S-shape. The central domain, comprised of five-helices, provides the subunit contacts in the functionally active tetramer. The active sites are located in clefts between the subunits and structural and mutagenic studies have identified several of the active site functional groups. While the catalytic activity of this enzyme has been known for nearly 100 years, a number of recent studies have revealed some interesting and unexpected new properties of this reasonably well-characterized enzyme. The non-linear kinetics that are seen under certain conditions have been shown to be caused by the presence of a separate regulatory site. The substrate, aspartic acid, can also play the role of an activator, binding at this site along with a required divalent metal ion. Truncation of the carboxyl terminus of aspartase at specific positions leads to an enhancement of the catalytic activity of the enzyme. Truncations in this region also have been found to introduce a new, non-enzymatic biological activity into aspartase, the ability to specifically enhance the activation of plasminogen to plasmin by tissue plasminogen activator. Even after a century of investigation there are clearly a number of aspects of this multifaceted enzyme that remain to be explored.

Aspartate aminotransferase activity is required for aspartate catabolism and symbiotic nitrogen fixation in Rhizobium meliloti.

Abstract: A mutant of Rhizobium meliloti, 4R3, which is unable to grow on aspartate has been isolated. The defect is specific to aspartate utilization, since 4R3 is not an auxotroph and grows as well as its parent strain on other carbon and nitrogen sources. The defect was correlated with an inability to fix nitrogen within nodules formed on alfalfa. Transport of aspartate into the mutant cells was found to be normal. Analysis of enzymes involved in aspartate catabolism showed a significantly lower level of aspartate aminotransferase activity in cell extracts of 4R3 than in the wild type. Two unrelated regions identified from a genomic cosmid bank each complemented the aspartate catabolism and symbiotic defects in 4R3. One of the cosmids was found to encode an aspartate aminotransferase enzyme and resulted in restoration of aspartate aminotransferase activity in the mutant. Analysis of the region cloned in this cosmid by transposon mutagenesis showed that mutations within this region generate the original mutant phenotypes. The second type of cosmid was found to encode an aromatic aminotransferase enzyme and resulted in highly elevated levels of aromatic aminotransferase activity. This enzyme apparently compensated for the mutation by its ability to partially utilize aspartate as a substrate. These findings demonstrate that R. meliloti contains an aspartate aminotransferase activity required for symbiotic nitrogen fixation and implicate aspartate as an essential substrate for bacteria in the nodule.