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P A Recsei

Publications and source records attributed to P A Recsei.

13 recordsLinked to original sources
PubMed1987

Cloning, sequence, and expression of the lysostaphin gene from Staphylococcus simulans.

A 1.5-kilobase-pair fragment of DNA that contains the lysostaphin gene from Staphylococcus simulans and its flanking sequences has been cloned and completely sequenced. The gene encodes a preproenzyme of Mr 42,000. The NH2-terminal sequence of the preproenzyme is composed of a signal peptide followed by seven tandem repeats of a 13-amino acid sequence. Conversion of prolysostaphin to the mature enzyme occurs extracellularly in cultures of S. simulans and involves removal of the NH2-terminal portion of the proenzyme that contains the tandem repeats. The high degree of homology of the repeats suggests that they have arisen by duplication of a 39-base-pair sequence of DNA. In S. simulans, the lysostaphin gene is present on a large beta-lactamase plasmid.

Amino Acid Sequence↗
PubMed1985

Pyruvoyl-dependent histidine decarboxylases. Mechanism of cleavage of the proenzyme from Lactobacillus buchneri.

When Lactobacillus buchneri was grown in the presence of [hydroxyl-18O]serine and pyridoxamine, no 18O was found in its histidine decarboxylase (HisDCase). However, when pyridoxamine was omitted from the growth medium, the labeled serine was incorporated into the HisDCase without dilution. Internal serine residues of the enzyme contained 18O only in their hydroxyl group, while the COOH-terminal serine of the beta chain of HisDCase contained equal amounts of 18O in both its hydroxyl and carboxyl group. This enzyme, like the HisDCase from Lactobacillus 30a (Recsei, P. A., Huynh, Q. K., and Snell, E. E. (1983) Proc. Natl. Acad. Sci. U.S.A. 80, 973-977), therefore, arises by nonhydrolytic serinolysis of its proenzyme. This result, together with comparative sequence data (Huynh, Q. K., and Snell, E. E. (1985) J. Biol. Chem. 260, 2798-2803), makes it highly probable that all of the pyruvoyl-dependent HisDCases arise by a similar mechanism from inactive proenzymes.

Amino Acid Sequence↗
PubMed1984

Histidine decarboxylase of Lactobacillus 30a. Sequences of the cyanogen bromide peptides from the alpha chain.

The alpha chain of histidine decarboxylase contains eight internal methionine residues. After reductive amination to convert the NH2-terminal pyruvoyl residue to an alanyl residue and cyanogen bromide treatment, 13 pure peptides were isolated. Four of these are incomplete cleavage products. The sequence of each of the remaining nine peptides was established by automated and manual degradation of the intact peptides and of smaller peptides obtained from tryptic, chymotryptic, and staphylococcal protease digests of the cyanogen bromide peptides. These results, together with the data on overlapping peptides reported in the accompanying paper (Huynh, Q. K., Recsei, P. A., Vaaler, G. L., and Snell, E. E. (1984) J. Biol. Chem. 259, 2833-2839), establish the complete amino acid sequence of the alpha chain.

Amino Acid Sequence↗
PubMed1984

Histidine decarboxylase of Lactobacillus 30a. Sequences of the overlapping peptides, the complete alpha chain, and prohistidine decarboxylase.

The complete amino acid sequence of the alpha chain of histidine decarboxylase of Lactobacillus 30a has been established by isolation and analysis of the eight methionine-containing tryptic peptides of this chain. These peptides provide the overlaps required to order all nine peptides derived by complete cyanogen bromide cleavage of the alpha chain (Huynh, Q.K., Vaaler, G.L., Recsei, P.A., and Snell, E.E. (1984) J. Biol. Chem. 259, 2826-2832). Ordering of six of the latter peptides was confirmed by isolation and analysis of four peptides derived by incomplete cyanogen bromide cleavage. The alpha chain is composed of 226 residues and has a molecular weight of 24,892 calculated from the sequence. These results and the previously determined sequence of the beta chain (Vaaler, G.L., Recsei, P.A., Fox, J.L., and Snell, E.E. (1982) J. Biol. Chem. 257, 12770-12774) establish the complete amino acid sequence of the enzyme and of the pi chain of prohistidine decarboxylase. The latter is composed of 307 amino acids and has a calculated molecular weight of 33,731. Four segments of the pi chain sequence are repeated. The bond between Ser-81 and Ser-82 that is cleaved during proenzyme activation is in an uncharged portion of the sequence that is rich in serine and threonine residues and is predicted to be part of a beta sheet structure.

Amino Acid Sequence↗
PubMed1984

Pyruvoyl enzymes.

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Adenosylmethionine Decarboxylase↗
PubMed1983

Pyruvoyl-dependent histidine decarboxylases from Clostridium perfringens and Lactobacillus buchneri. Comparative structures and properties.

Histidine decarboxylase (EC 4.1.1.22) was purified to homogeneity from Clostridium perfringens and also from Lactobacillus buchneri. Both enzymes are composed of alpha and beta subunits, with an essential pyruvoyl group bound to the alpha subunit. In this respect and also in molecular weight of both the alpha and beta subunits and the native enzyme, they closely resemble the previously described (Riley, W.D., and Snell, E. E. (1970) Biochemistry 9, 1485-1491) histidine decarboxylase from Lactobacillus 30a. Rabbit antibodies to the latter enzyme cross-react incompletely with the decarboxylase from L. buchneri but not with that from C. perfringens in double diffusion tests. The clostridial decarboxylase differs substantially from the Lactobacillus 30a enzyme in amino acid composition and, unlike the latter enzyme, requires high ionic strength (I approximately 1.4 M) for maximum activity. The enzymes also differ in rates of electrophoretic migration. A proenzyme for the decarboxylase similar to that previously found in Lactobacillus 30a was detected in immunoprecipitates of extracts of L. buchneri. We conclude that these proteins arise from pyruvate-free precursor proteins by similar mechanisms and probably have diverged from a common ancestral protein.

Amino Acids↗
PubMed1983

Conversion of prohistidine decarboxylase to histidine decarboxylase: peptide chain cleavage by nonhydrolytic serinolysis.

Unlabeled prohistidine decarboxylase and prohistidine decarboxylase containing L-[carboxyl-(18)O]serine or L-[hydroxyl-(18)O]serine were isolated in homogeneous form from mutant 3 of Lactobacillus 30a grown with the appropriately labeled serine. There was no randomization or redistribution of label during growth, isolation of the protein, or enzymatic hydrolysis and reisolation of the labeled amino acids. These proteins were used to show that during proenzyme activation, in which individual pi subunits of the proenzyme are converted to alpha and beta subunits of the active enzyme [Formula: see text] (in which pi, alpha, and beta subunits have the partial structures shown and Prv designates a pyruvoyl group), no (18)O from H(2) (18)O is incorporated into the newly formed carboxyl terminus (Ser-81) of the beta chain, although no labilization of (18)O from proenzyme labeled with L-[carboxyl-(18)O]serine occurred when the proenzyme was activated in H(2) (16)O by the same procedures. The additional oxygen atom present in the carboxyl group of Ser-81 of the beta subunit is transferred from the hydroxyl group of Ser-82 of the proenzyme during the activation reaction. The same result was obtained with wild-type enzyme formed intracellularly. Peptide bond cleavage during activation of the proenzyme thus proceeds by a hitherto unobserved direct or indirect "serinolysis" coupled to alpha,beta-elimination at Ser-82 to yield the pyruvoyl group of the alpha subunit, rather than by hydrolysis. Possible mechanisms for the reaction are discussed briefly.

Amino Acid Sequence↗
PubMed1982

Histidine decarboxylase of Lactobacillus 30a. Comparative sequences of the beta chain from wild type and mutant enzymes.

Manual and automated sequencing of peptides isolated from tryptic, chymotryptic, and Staphylococcus aureus V8 protease digests of the beta chain of histidine decarboxylase from Lactobacillus 30a have established the following sequence for the wild type protein: NH2-Ser-Gly-Leu-Asp-Ala-Lys-Leu-Asn-Lys-Leu-Gly-Val-Asp-Arg-Ile-Ala-Ile-Ser-Pro -Tyr-Lys-Gln-Trp-Thr-Arg-Gly-Tyr-Met-Glu-Pro-Gly-Asn-Ile-Gly-Asn-Gly-Tyr-Val-Thr-Gly-Leu-Lys-Val-Asp-Ala-Gly-Val-Arg-Asp-Lys-Ser-Asp-Asp-Asp-Val-Leu-Asp-Gly-I le-Val-Ser-Tyr-Asp-Arg-Ala-Glu-Thr-Lys-Asn-Ala-Tyr-Ile-Gly-Gln-Ile-Asn-Met-Thr- Thr-Ala-Ser-COOH The beta chain from mutant 3 of this organism shows two amino acid replacements: Ser-51 is replaced by Ala and Gly-58 by Asp. These amino acid replacements result in a significant increase in the predicted alpha-helical content and a significant decrease in the isoelectric point of the mutant beta chain and are consistent with changes in physical and catalytic properties of the mutant histidine decarboxylase. In addition, about 15% of the mutant chains contain Met-Ser at the NH2 terminus rather than Ser. Asn-35 is partially deamidated in both proteins. Structural comparisons show that the histidine decarboxylase from Lactobacillus 30a and a similar pyruvoyl enzyme from Micrococcus sp. n. have evolved from a common ancestral protein.

Amino Acid Sequence↗
PubMed1981

Crystallization and subunit structure of histidine decarboxylase from Lactobacillus 30a.

Histidine decarboxylase from Lactobacillus 30a has been crystallized in a variety of forms which together indicate a revised subunit structure for the native particle. Octahedral crystals of the wild type enzyme obtained at room temperature from ammonium sulfate solutions in microdiffusion cells belong to tetragonal space group I4122 with a = b = 222 A and c = 107.5 A. Trigonal and hexagonal plates of prohistidine decarboxylase and activated proenzyme obtained at 4 degrees C from polyethyleneglycol solutions by vapor equilibration using the hanging drop technique belong to the trigonal space group P321 with a = b = 100 A and c = 164 A. The space group symmetries and unit cell contents of these crystals indicate 32 point group symmetry for the subunit structure of these enzymes. Sedimentation coefficients of wild type enzyme measured as a function of ionic strength at pH 7.0 indicate a rapid equilibrium between species varying from 6.9 S to 9.4 S. Sedimentation equilibrium analysis demonstrated the existence of a nearly homogeneous particle with Mr congruent to 208,000 at ionic strengths above I = 0.20, while an additional species of approximately one-half that molecular weight is observed at very ionic strengths (I = 0.2). At the pH optimum of the enzyme (pH 4.8), te larger species is dominant at all ionic strengths tested. Electron micrographs of native wild type enzyme show a dominant tetrahedral particle approximately 60 A on an edge while similar micrographs of enzyme cross-linked with glutaraldehyde show a dumbbell-shaped particle approximately 60 A in width and 120 A in length. These results establish that: (a) the native enzyme has a Mr congruent to 208,000 and a subunit composition (alpha beta)6; (b) the proenzyme has a subunit composition (pi)6; and (c) stable (alpha beta)3 and (pi) 3 particles exist under certain conditions.

Carboxy-Lyases↗
PubMed1972

Histidine decarboxylaseless mutants of Lactobacillus 30a: isolation and growth properties.

Mutants of Lactobacillus 30a deficient in their ability to form an inducible histidine decarboxylase (EC 4.1.1.22) were selected by plating nitrosoguanidine-treated cultures on a medium containing histidine and methyl red. Wild-type organisms produce histamine, thus raising the pH and forming yellow colonies; mutant colonies remain red. In the presence of added histidine, decarboxylase-producing cultures grow more heavily than mutant cultures when the initial pH of the growth medium is low or when the lactic acid produced lowers the pH to growth-limiting values. Addition of the decarboxylation products, histamine and carbon dioxide, did not favor growth in crude medium.

Azo Compounds↗