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

The nucleotide sequence has been determined of a 1400 bp fragment from the chromosome of Yersinia enterocolitica containing the gene for beta-lactamase I. An ORF of 882 bp was identified, which could code for a polypeptide of 294 amino acids, closely related to other beta-lactamases of molecular class A. Amino acids 1-30 could constitute a signal peptide. The mature protein would be 264 amino acids long with a calculated pI of 6.2. Alignment of the amino acid sequence of the class A beta-lactamases suggested the existence of two subgroups in the same class, and this is discussed in the context of the evolution of the enzymes.
Mol Gen Genet 1991 Aug
PMID:Nucleotide sequence of a new class A beta-lactamase gene from the chromosome of Yersinia enterocolitica: implications for the evolution of class A beta-lactamases. 188 8

The effects of 25-fold overproduction of Escherichia coli signal peptidase I (SPase I) on the processing kinetics of various (hybrid) secretory proteins, comprising fusions between signal sequence functions selected from the Bacillus subtilis chromosome and the mature part of TEM-beta-lactamase, were studied in E. coli. One precursor (pre[A2d]-beta-lactamase) showed an enhanced processing rate, and consequently, a highly improved release of the mature enzyme into the periplasm. A minor fraction of a second hybrid precursor (pre[A13i]-beta-lactamase), which was not processed under standard conditions of SPase I synthesis, was shown to be processed under conditions of SPase I overproduction. However, this did not result in efficient release of the mature beta-lactamase into the periplasm. In contrast, the processing rates of wild-type pre-beta-lactamase and pre(A2)-beta-lactamase, already high under standard conditions, were not detectably altered by SPase I overproduction. These results demonstrate that the availability of SPase I can be a limiting factor in protein export in E. coli, in particular with respect to (hybrid) precursor proteins showing low (SPase I) processing efficiencies.
Mol Gen Genet 1991 May
PMID:Signal peptidase I overproduction results in increased efficiencies of export and maturation of hybrid secretory proteins in Escherichia coli. 190 37

We have developed a 33 mer probe that hybridizes to the serine active site of the chromosomal ampC beta-lactamase gene of Pseudomonas aeruginosa and the Enterobacteriaceae. We tested this probe against a variety of Enterobacteriaceae, and a series of 23 P. aeruginosa by dot-blots and selected Southern blots. This probe is an alternative or supplement to enzyme studies for characterizing the class of a Gram-negative rod's beta-lactamase and is a useful tool for studies of Pseudomonas beta-lactamase regulation.
Mol Cell Probes 1991 Apr
PMID:A general ampC active site oligonucleotide probe for gram-negative rods. 190 78

AmpR, the transcriptional regulator for the Citrobacter freundii ampC beta-lactamase gene, was purified. The purified AmpR had DNA-binding activity, the same molecular mass (32 kDa) on sodium dodecyl sulphate/polyacrylamide gel electrophoresis as previously described, and N-terminal sequencing of the first 15 amino acids was in agreement with that predicted from the nucleotide sequence. Two mutants were isolated that abolish DNA-binding and beta-lactamase induction and which map in the amino- and carboxyl-terminal ends of AmpR, respectively. The mutation in the amino terminus (S35F) was located in a helix-turn-helix region showing high homology to other members of the LysR regulator family. Therefore this mutation may directly abolish the contact between AmpR and its operator sequence. It is suggested that the C-terminal mutation (Y264N) affects subunit interactions in AmpR. One constitutive mutant was isolated which mapped in the centre of the ampR gene. This G102E mutant leads to constitutive beta-lactamase expression in the absence of both beta-lactam inducer and ampG, a gene essential for induction in wild-type enterobacteria. Another mutant protein, D135Y, showed wild-type properties in an ampG+ and an ampG::kan background, but could, unlike wild-type AmpR, activate the ampC gene in an ampG1 mutant background. It is thought that ampG1 is a missense mutant. These two types of ampR mutants suggest that activation of ampC transcription is dependent on the conversion of AmpR into a transcriptional activator and that this activation may normally involve interactions with AmpG.
Mol Microbiol 1991 Jul
PMID:Purification and mutant analysis of Citrobacter freundii AmpR, the regulator for chromosomal AmpC beta-lactamase. 194 5

The class B, metallo-beta-lactamase genes ccrA (carbapenem- and cephamycin resistance) from three Bacteroides fragilis isolates--QMCN3, QMCN4, and TAL3636--were cloned and expressed in Escherichia coli. Cloning of the genes, by selecting for ampicillin resistance, was facilitated by two classes of Escherichia coli chromosomal mutations which resulted in at least a 5-10-fold increase in metallo-beta-lactamase enzymatic activity. The observed increase in enzymatic activity is due to either increased translation of the ccrA gene or an effect on localization or stability of the protein. Comparison of the DNA sequences of the three ccrA genes revealed that their protein-coding sequences shared greater than 97% DNA sequence identity. However, the 5' upstream sequence for the TAL3636 ccrA gene was unrelated to that of the other two genes.
Mol Microbiol 1991 May
PMID:Escherichia coli chromosomal mutations that permit direct cloning of the Bacteroides fragilis metallo-beta-lactamase gene, ccrA. 195 98

Haemolysin B (HlyB) is essential for secretion of the 107 x 10(3) Mr haemolysin A protein from Escherichia coli and is a member of a family of highly conserved, apparently ATP-dependent surface proteins in many organisms. We have shown in this study that both HlyB and HlyD fractionate primarily with the cytoplasmic membrane of E. coli and are accessible to proteases after removal of the outer membrane. We have measured experimentally the topological organization of HlyB within the membrane by construction of fusions to beta-lactamase as a reporter. The predicted folding of HlyB, with a minimum of six transmembrane segments, does not always coincide with regions of highest average hydrophobicity. This suggests that HlyB may have a novel organization within the bilayer. From our data and comparative sequence analysis, we have been able to predict very similar topological models for the other members of the HlyB family.
J Mol Biol 1991 Feb 05
PMID:Analysis of the membrane organization of an Escherichia coli protein translocator, HlyB, a member of a large family of prokaryote and eukaryote surface transport proteins. 199 34

The crystal structure of a class A beta-lactamase from Staphylococcus aureus PC1 has been refined at 2.0 A resolution. The resulting crystallographic R-factor (R = sigma h parallel Fo[-]Fc parallel/sigma h[Fo], where [Fo] and [Fc] are the observed and calculated structure factor amplitudes, respectively), is 0.163 for the 17,547 reflections with I greater than or equal to 2 sigma (I) within the 8.0 A to 2.0 A resolution range. The molecule consists of two closely associated domains. One domain is formed by a five-stranded antiparallel beta-sheet with three helices packing against a face of the sheet. The second domain is formed mostly by helices that pack against the second face of the sheet. The active site is located in the interface between the two domains, and many of the residues that form it are conserved in all known sequences of class A beta-lactamases. Similar to the serine proteases, an oxyanion hole is implicated in catalysis. It is formed by two main-chain nitrogen atoms, that of the catalytic seryl residue, Ser70, and that of Gln237 on an edge beta-strand of the major beta-sheet. Ser70 is interacting with another conserved seryl residue, Ser130, located between the two ammonium groups of the functionally important lysine residues, Lys73 and Lys234. Such intricate interactions point to a possible catalytic role for this second seryl residue. Another key catalytic residue is Glu166. There are several unusual structural features associated with the active site. (1) A cis peptide bond has been identified between the catalytic Glu166 and Ile167. (2) Ala69 and Leu220 have strained phi, psi dihedral angles making close contacts that restrict the conformation of the active site beta-strand involved in the formation of the oxyanion hole. (3) A buried aspartate residue, the conserved Asp233, is located next to the active site Lys234. It is interacting with another buried aspartyl residue, Asp246. An internal solvent molecule is also involved, but the rest of its interactions with the protein indicate it is not a cation. (4) Another conserved aspartyl residue that is desolvated is Asp131, adjacent to Ser130. Its charge is stabilized by interactions with four main-chain nitrogen atoms. (5) An internal cavity underneath the active site depression is filled with six solvent molecules. This, and an adjacent cavity occupied by three solvent molecules partially separate the omega-loop associated with the active site from the rest of the protein.(ABSTRACT TRUNCATED AT 250 WORDS)
J Mol Biol 1991 Feb 20
PMID:Refined crystal structure of beta-lactamase from Staphylococcus aureus PC1 at 2.0 A resolution. 200 20

While several proteins, including beta-lactamase, cytochrome c and apomyoglobin, are maximally unfolded at pH 2 by HCl in the absence of salt, the addition of anions, either from salt or acid, co-operatively induces the unfolded proteins to refold to a molten globule state, because anions bind preferentially to the compact molten globule state compared to the extended unfolded state. To study the role of the anion-dependent conformational transition at neutral pH, we synthesized a model polypeptide of 51 amino acid residues, consisting of tandem repeats of a Lys-Lys-Leu-Leu sequence and containing a turn sequence, Asn-Pro-Gly, at the center of the molecule. The model polypeptide showed no significant conformation by circular dichroism under conditions of low salt at neutral pH. However, addition of anions, either from salt or acid, induced the folding transition to an alpha-helical conformational state. The order of effectiveness of various anions in inducing the folding transition was consistent with the series of anions in inducing the molten globule of the acid-denatured protein. This suggests that the helical state of the model polypeptide is equivalent to the molten globule state. At pH values above 9, the model polypeptide also took an alpha-helical conformation, which was very similar to that induced by anions. On the basis of the chloride and pH-dependent conformational transitions, a phase diagram for the conformational states was constructed. The phase diagram was explained simply by assuming that the conformational transition is linked to the proton and the anion bindings to a limited number of amino groups and that anions bind only to the protonated groups.
J Mol Biol 1991 Mar 20
PMID:Anion and pH-dependent conformational transition of an amphiphilic polypeptide. 201 Sep 16

The rate of folding of the precursor of beta-lactamase is not influenced by the presence of SecB under conditions in which GroEL/ES retards the folding. Wild-type beta-lactamase and several mutants in the signal or the mature protein, affecting either transport or enzyme kinetics and probably folding, were examined for total expression, total enzymatic activity, and transported beta-lactamase (in vivo resistance) in secB- and secB+ strains. We conclude that there is no indication of any relevant interaction between SecB and pre-beta-lactamase in vitro, nor did the secB- mutation affect the transport of wild-type beta-lactamase or any of the mutant in vivo. Thus, putative Escherichia coli "folding modulators' must be of limited specificity.
Mol Microbiol 1991 Jan
PMID:Folding in vitro and transport in vivo of pre-beta-lactamase are SecB independent. 201 98

Gene fusions were constructed between Ste2, the receptor for the Saccharomyces cerevisiae alpha-factor, and beta la, the secreted form of beta-lactamase encoded by the bla gene of pBR322. The Ste2 and beta la components were linked by a processing fragment (P) from the yeast killer preprotoxin containing a C-terminal lysine-arginine site for cleavage by the Golgi-associated Kex2 protease. Ste2 is predicted to have a rhodopsinlike topology, with an external N terminus and seven transmembrane segments. Fusions to three of the four Ste2 domains predicted to be external resulted in beta la secretion from yeast cells. A fusion at a site just preceding the first transmembrane segment was an exception; the product was cell associated, indicating that the first 44 residues of Ste2 are insufficient to direct secretion of beta la; translocation of this domain presumably requires the downstream transmembrane segment. Expression of fusions located in two domains predicted to be cytoplasmic failed to result in beta la secretion. Following insertion of the preprotoxin signal peptide (S) between the Ste2 and P components of these cytoplasmic fusions, secretion of beta la activity occurred, which is consistent with inversion of the orientation of the beta la reporter. Conversely, insertion of S between Ste2 and P in an external fusion sharply reduced beta la secretion. Complementary information about both cytoplasmic and external domains of Ste2 was therefore provided, and most aspects of the predicted topology were confirmed. The steady-state levels of beta la detected were low, presumably because of efficient degradation of the fusions in the secretory pathway; levels, however, were easily detectable. This method should be valuable in the analysis of in vivo topologies of both homologous and foreign plasma membrane proteins expressed in yeast cells.
Mol Cell Biol 1991 May
PMID:In vivo topological analysis of Ste2, a yeast plasma membrane protein, by using beta-lactamase gene fusions. 201 68


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