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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A genomic library of Mycobacterium bovis BCG has been constructed by cloning DNA partially digested with Sau3A into the Escherichia coli expression vector pAS1. The gene coding for ornithine carbamoyl-transferase (EC.2.1.3.3; OTCase), hereafter referred to as argF, was isolated from the library by complementation of a double argF-argI mutant of E. coli and its sequence was determined. The translation initiation codon used, GTG, was identified by comparing the amino acid sequence deduced from the gene with the N-terminal sequence of the corresponding purified protein. On this basis, the M. bovis BCG OTCase monomer consists of 307 amino acid residues and displays about 44% identity with other OTCases, the most closely related homologue being the anabolic enzyme of Pseudomonas aeruginosa. The native enzyme has an estimated molecular mass of 110 kDa, suggesting a trimeric structure as is the case for most of the anabolic OTCases known from various organisms.
Mol Gen Genet 1992 Sep
PMID:Molecular cloning, characterization and purification of ornithine carbamoyltransferase from Mycobacterium bovis BCG. 140 93

The effect of arginine on the growth and metabolism of Giardia intestinalis trophozoites was determined. Supplementation of the normal growth medium (Diamond's TYI-S-33) with 5 or 10 mM arginine accelerated trophozoite growth over the first 2 days. There was a corresponding rapid utilisation of arginine, with none being detectable after this time. The decrease was associated with the appearance in the growth medium of 1 mol of ornithine and 2 mol of ammonia per mol of arginine utilised, the stoichiometry being consistent with the operation of the arginine dihydrolase pathway. Subsequently, there was a decrease in the ammonia concentration in the medium. Removal of arginine from the medium by pretreatment with arginase substantially decreased cell growth. In TYI-S-33 medium containing no added glucose, instead of the normal 50 mM glucose concentration, arginine supplementation also increased cell growth over the first 2 days, with concurrent stoichiometric production of ornithine and ammonia. However, in these conditions, the ammonia concentration remained elevated. This suggests that under normal conditions there is re-uptake of ammonia, which is glucose dependent. The observations confirm the operation of a functional arginine dihydrolase pathway in G. intestinalis. The concordance of cessation of rapid growth with the depletion of arginine, and the beneficial effect on growth of arginine supplementation suggests that arginine availability is a limiting factor during the initial stages of rapid growth. It would appear that arginine is a major potential energy source during the initial stages of giardial growth, and that supplementation of Diamond's TYI-S-33 medium with additional arginine may provide an improved in vitro culture medium.
Mol Biochem Parasitol 1992 Jul
PMID:Arginine metabolism during culture of Giardia intestinalis. 150 49

The nucleotide sequence required for a fully functional promoter and operator of the Pseudomonas aeruginosa argF gene (argFpo), the arginine-repressible gene for anabolic ornithine carbamoyltransferase, was defined within a 160 bp region. The streptomycin (Sm) resistance genes strAB of plasmid RSF1010 were fused to argFpo. This construct in P. aeruginosa strain PAO conferred resistance to Sm. Mutants of strain PAO were selected which were resistant to Sm in the presence of arginine due to constitutive expression of argFpo-strAB. These mutants were designated argR. They were unable to grow or grew poorly on arginine or ornithine as the sole carbon and nitrogen source. This growth defect (Aru-/Oru- phenotype) was correlated with a reduced level of N-succinylornithine aminotransferase, an enzyme participating in the major aerobic pathway for arginine and ornithine catabolism in this organism. The argR mutants were classified into four groups by transduction analysis and three argR mutations were mapped on the PAO chromosome. argR9901 and argR9902 were co-transducible with car-9 (at 1 min) and thus close to the oru-310 locus; argR9906 was localized in the oruI (= aru) gene cluster (67 min). Some aru mutants, which have been isolated previously and which produce very low amounts of all enzymes in the arginine succinyltransferase pathway, were unable to repress the argF gene in an arginine medium. Thus, P. aeruginosa PAO appears to have multiple genes that are involved in the regulation of both the anabolic argF and the catabolic aru genes.
Mol Gen Genet 1992 Feb
PMID:Mutations affecting regulation of the anabolic argF and the catabolic aru genes in Pseudomonas aeruginosa PAO. 153 97

The entire nucleotide sequence of the Bacillus brevis grsB gene encoding the gramicidin S synthetase 2, which activates and condenses the four amino acids proline, valine, ornithine and leucine has been determined. The gene contains an open reading frame of 13,359 bp which encodes a protein of 4453 amino acids with a predicted Mr of 510,287. The gene is located within the gramicidin S biosynthetic operon, also containing the genes grsT and grsA, whose nucleotide sequences have been determined previously. Within the GrsB amino acid sequence four conserved and repeated domains of about 600 amino acids (45-50% identity) have been identified. The four domains are separated by non-homologous sequences of about 500 amino acids. The domains also share a high degree of similarity (20-70%) with eight peptide synthetases of bacterial and fungal origin as well as with conserved sequences of nine other adenylate-forming enzymes of diverse origin. On the basis of sequence homology and functional similarities, we infer that those enzymes share a common evolutionary origin and present a phylogenetic tree for this superfamily of domain-bearing enzymes.
Mol Microbiol 1992 Feb
PMID:Four homologous domains in the primary structure of GrsB are related to domains in a superfamily of adenylate-forming enzymes. 144 81

The periplasmic binding protein LAO from Salmonella typhimurium, which is involved in lysine, arginine and ornithine transport, has been crystallized together with one of its ligands, arginine (LAO-Arg). Preliminary X-ray diffraction studies of LAO-Arg crystal show that it belongs to the orthorhombic space group P2(1)2(1)2(1) and has the unit cell dimensions of a = 37.65 A, b = 59.45 A, c = 115.91 A. Crystals of the LAO-Arg complex diffract beyond 2.0 A resolution.
J Mol Biol 1992 Jun 20
PMID:Crystallization and preliminary X-ray studies of the liganded lysine, arginine, ornithine-binding protein from Salmonella typhimurium. 161 94

Two Corynebacterium glutamicum mutants defective in lysine uptake were identified by analysing mutants resistant to S-(2-aminoethyl)-cysteine (AEC). A 5.6 kb genomic DNA fragment restoring AEC sensitivity and lysine uptake was isolated. A 4.2 kb subfragment was sequenced and three open reading frames were identified. Subcloning and gene disruption experiments showed that only the first open reading frame, termed lysl, is involved in lysine uptake. Lysl consists of 501 amino acids with a Mr of 53600. The hydrophobicity profile suggests that the lysl gene product is an integral membrane protein with 13 transmembrane segments. The amino acid sequence of lysl displays strong homology to that of the arcD gene product of Pseudomonas aeruginosa, which is proposed to act as an arginine-ornithine antiporter. Investigation of the influence of the lysl gene on lysine secretion suggests the existence of a separate lysine efflux system in C. glutamicum.
Mol Microbiol 1991 Dec
PMID:Molecular analysis of the Corynebacterium glutamicum lysl gene involved in lysine uptake. 166 21

Polyamine levels were measured by means of high-performance liquid chromatography in Langendorff-perfused rat hearts subjected to the calcium paradox protocol. The concentrations of putrescine, spermidine and spermine did not change significantly during calcium-free perfusion but decreased when calcium was readmitted. This decrease was due to membrane disruption and release of the polyamines into the coronary effluent. The sum of released and remaining spermidine exceeded the concentration of spermidine in control hearts, but, for spermine, this sum was lower than the control level. The addition of 0.5 mM EGTA to the calcium-free solution raised the myocardial concentrations of putrescine and spermidine and enhanced the net increase of spermidine on calcium repletion. DL-alpha-Difluoromethylornithine (DFMO) inhibited these increases and lowered the putrescine level during all perfusion stages. External polyamines had a negative inotropic effect and inhibited the loss of myoglobin on calcium repletion (order of effectiveness: spermine greater than spermine greater than putrescine). Inhibition of contractions by the combined action of verapamil and ryanodine or by potassium depolarization did not prevent myoglobin loss. External polyamines had no effect on high K/low Na contractures, which were mediated mainly by Na-Ca exchange. Calcium-free perfusion in the presence of 0.5 to 1 mM EGTA improved the membrane protection by polyamines or by diamines and analogues, like ornithine, 1,3-diaminopropane, or DFMO, which, in the absence of EGTA, gave no clear protection. It is concluded that calcium depletion and repletion influences myocardiaal polyamine concentrations by (1) membrane disruption and release of polyamines into the coronary effluent, and (2) probably by a stimulation of ornithine decarboxylase activity. The changes in polyamine concentrations do not seem to have any causal role in calcium overload and cell death. Exogenous polyamines protect against membrane damage.
J Mol Cell Cardiol 1991 Mar
PMID:Polyamines and the calcium paradox in rat hearts. 190 6

Inhibition of tryptophanase-catalyzed decomposition of S-(o-nitrophenyl)-L-cysteine by a variety of amino acids has been investigated. For amino acids similar to the natural substrate and for those having minimal steric requirements for the side chain, the linear correlation exists between-RTlnKi and side chain hydrophobicity. L-ornithine and L-arginine are anomalously potent inhibitors taking into account low hydrophobicity of their side chains. This can be explained by an interaction between a positively charged group of the side chain of L-arginine or L-ornithine and a nucleophilic group of the active site. The comparison of affinity of tryptophanase for L-phenylalanine and L-homophenylalanine indicates that there is a special locus in the active site where aromatic groups are bound and oriented approximately parallel to the cofactor plane experiencing no steric hindrance. For a large number of amino acids the rates of the enzymic alpha-proton exchange in 2H2O are comparable with the rate of the reaction with L-tryptophan. Very low rate of alpha-proton exchange observed with L-alanine is an exception.
Mol Biol (Mosk)
PMID:[Factors, determining the effectiveness of tryptophanase interaction with amino acids]. 194 57

Our previous studies suggested that the ornithine decarboxylase inhibitor alpha-difluoromethylornithine (DFMO) inhibits bone resorption by mechanisms that are independent of polyamine depletion. To determine whether DFMO prevents calcitriol-stimulated bone resorption by acting at a step before or after osteoclast activation, we compared the effects of DFMO on release of calcium and beta-glucuronidase from cultured neonatal mouse calvaria. DFMO, at concentrations of 7.5-20 mM, inhibited release of calcium from calcitriol-stimulated calvaria but failed to inhibit the calcitriol-stimulated increase in beta-glucuronidase secretion. In contrast, ornithine, putrescine, spermidine, and spermine, at concentrations with effects on resorption comparable to those of DFMO, inhibited the effects of calcitriol on both calcium and beta-glucuronidase release. NaF (0.2 mM), like DFMO, inhibited calcitriol-stimulated calcium release without affecting medium beta-glucuronidase activity, whereas elevated phosphate (3 mM) inhibited both activities. The results suggest that DFMO, over the concentration range studied, inhibits calcium release by making the matrix resistant to resorption rather than by acting at a cellular locus.
Mol Pharmacol 1991 Apr
PMID:Alpha-difluoromethylornithine inhibits bone resorption in vitro without decreasing beta-glucuronidase release. 201 55

Ornithine transcarbamoylase catalyzes the formation of L-citrulline from carbamoyl phosphate and L-ornithine. The anabolic enzyme from Escherichia coli is composed of three identical subunits and resembles, in both primary and quaternary structures, the catalytic unit of aspartate transcarbamoylase. However, ornithine transcarbamoylase has no regulatory subunits. Although this enzyme does not bind its substrates co-operatively, fluorescence spectroscopic experiments show that zinc adds allosterically to the free enzyme. The metal binding process is dictated by deprotonation of an enzymic group with a pKa less than 7. The saturation binding curve of the metal ion is sigmoidal and yields a Hill coefficient of 1.6 at pH 8.5 and 25 degrees C. In the absence of substrates, zinc further promotes a slow enzyme isomerization, which occurs with a first-order rate constant of 7 min-1; thus, the metal is a slow, tight-binding inhibitor. The isomerized enzyme is inactive and contains three zinc ions. When the enzyme is first bound with carbamoyl phosphate, steady-state kinetic assays reveal that zinc again binds co-operatively to the binary enzyme complex, with a Hill coefficient of 1.5, but the metal ion now behaves simply as a classical, reversible inhibitor; it is competitive against L-ornithine, non-competitive against carbamoyl phosphate, and induces no enzyme isomerization. However, as a result of the competition between zinc and L-ornithine for the same site on the enzyme, the L-ornithine saturation curve becomes sigmoidal. Displacement of the allosteric zinc from the enzyme by L-ornithine is the cause of co-operative addition of this substrate. The combined results suggest that zinc regulates ornithine transcarbamoylase via two routes: (1) as an allosteric cofactor of the substrate-bound enzyme in mediating site-site interactions; and (2) as a slow, tight-binding inhibitor of the free enzyme in inducing inactivation. The concentration of zinc that is effective for action is in the micromolar range. The finding that E. coli ornithine transcarbamoylase can be induced to express co-operativity in binding its substrates has recently been confirmed by site-directed mutagenesis experiments. When the active site residue Arg106 is altered to a glycine, the resultant mutant enzyme exhibits both homotropic and heterotropic interactions towards its substrates. In view of the quaternary structure of holoenzyme aspartate transcarbamoylase, the "silent" co-operativity of ornithine transcarbamoylase is of particular interest in the study of evolution of complex, regulatory proteins.
J Mol Biol 1990 Jan 05
PMID:Zn2+ regulation of ornithine transcarbamoylase. I. Mechanism of action. 210 98


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