Close evolutionary relationship between the chromosomally encoded beta-lactamase gene of Klebsiella pneumoniae and the TEM beta-lactamase gene mediated by R plasmids

Sixty-three percent homology of nucleotide sequence and 67% homology of deduced amino acid sequence were found between the chromosomally encoded beta-lactamase gene of Klebsiella pneumoniae and the TEM beta-lactamase of transposon Tn3. Moreover, 22 out of 24 amino acid residues are identical around the predicted active site. It is therefore suggested that these two kinds of beta-lactamases share a common evolutionary origin. The 0.5 kb DNA fragment of the cloned gene hybridized specifically with the chromosomal DNA of all the K. pneumoniae strains tested which had been isolated in Japan, USA and Europe.     

Klebsiella oxytoca: Resistance to aztreonam by overproduction of the chromosomally encoded β-lactamase

Abstract Aztreonam-resistant Klebsiella oxytoca strain SL7811 was selected on agar containing 1 μg of aztreonam per ml from a susceptible strain SL781. The MICs for the resistant mutant towards penicillins, aztreonam and ceftriaxone were much higher, to cefotaxime slightly higher and to ceftazidime unchanged. Synthesis of β-lactamase was 223-fold greater in the mutant compared with the susceptible strain. SL781 and its resistant mutant SL7811 produced β-lactamase with the same isoelectric point and substrate profile. The β-lactamase genes from SL781 and SL7811 were cloned in plasmid pBGS18 giving pBOF-1 and pBOF-4 respectively. The sequences of the two putative promoters indicated two modifications in the resistant plasmid pBOF-4: a transversion (G → T) in the first base of the − 10 consensus sequence and a deletion of one C residue four base pairs upstream of the − 10 hexamer.     

The amino acid sequence of glutamate decarboxylase from Escherichia coli. Evolutionary relationship between mammalian and bacterial enzymes.

The amino acid sequence of glutamate decarboxylase from Escherichia coli was solved by a combination of automated Edman degradation of peptide fragments derived by proteolytic and chemical cleavage and sequencing of DNA. Correct alignment of three peptides, for which no peptide overlaps were available, was achieved by sequencing a 1.1-kbp fragment of DNA produced by a polymerase-chain reaction using primers corresponding to sequences known to be in amino-terminal and carboxy-terminal regions of the protein. Sequence similarity (24% identity) with mammalian glutamate decarboxylase was found to be limited to a 55-residue sequence around the lysine residue that binds the coenzyme. Stronger similarity (38% identity), again confined to the same region, is seen with bacterial pyridoxal-phosphate-dependent histidine decarboxylase.     

Single amino acid substitution between SHV-1 beta-lactamase and cefotaxime-hydrolyzing SHV-2 enzyme

SHV-2 beta-lactamase was purified from an overproducing variant of a clinical isolate of Escherichia coli resistant to cefotaxime. Pure protein was digested by trypsin and Lys-C endoproteinase. Proteolytic peptides, isolated by reverse-phase HPLC, were submitted to manual Edman degradation and aligned by homology with the sequence of SHV-1 beta-lactamase. A putative amino acid sequence was deduced. Structural comparison revealed that SHV-2 differed from SHV-1 by only one amino acid, Gly----Ser, at position 213 of the mature protein.     

Prokaryote-eukaryote relationship and the amino acid sequence of plastocyanin from Anabaena variabilis.

The amino acid sequence of plastocyanin from the prokaryotic blue-green alga Anabaena variabilis was determined. The protein consists of a single polypeptide chain of 105 residues. The amino acid sequence of the plastocyanin was compared with that of the eukaryotic green alga Chlorella fusca and with those of higher-plant plastocyanins. The considerable similarity between the prokaryotic and eukaryotic plastocyanins is discussed. Detailed evidence for the sequence of the protein has been deposited as Supplementary Publication SUP 50051 (13 pages) at the British Library (Lending Division), Boston Spa, Wetherby, W. Yorkshire LS23 7BQ, U.K., from whom copies may be obtained on the terms given in Biochem J. (1975) 145, 5.     

The partial amino acid sequence of the extracellular beta-lactamase I of Bacillus cereus 569/H.

The chemical structure of the extracellular beta-lactamase I of Bacillus cereus 569/H was investigated. Three electrophoretically homogenous charge variants of this enzyme were isolated and amino acid analysis of each revealed no significant differences. However, a degree of N-terminal heterogeneity was found by direct end-group modification of the protein and also on alignment of peptides from tryptic and chymotryptic digestion. The N-terminal heterogeneity observed was great enough to explain the production of the beta-lactamase I isoenzymes which are probably produced by postsynthesis modification of a single gene product. Over 80% of the amino acid sequence of beta-lactamase I was determined by the detailed analysis of peptides derived from tryptic, chymotryptic and thermolytic digests. Five polypeptide fragments were constructed from these data and aligned by comparison with the known amino acid sequences of the penicillinases produced by Bacillus licheniformis and Staphylococcus aureus (Ambler & Meadway, 1969). About 60% of the proposed sequence was identical with that of B. licheniformis penicillinase, whereas the S. aureus enzyme had only about 40% of its residues in common with beta-lactamase I. These results are discussed with reference to the possible evolutionary relationships existing between known beta-lactamases. Detailed evidence for the amino acid sequence proposed has been deposited as Supplementary Publication SUP 50044 (27 pages) at the British Library (Lending Division), Boston Spa, Wetherby, W. Yorks. LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1975), 145, 5.     

Nucleotide sequence of the Klebsiella pneumoniae nifD gene and predicted amino acid sequence of the alpha-subunit of nitrogenase MoFe protein.

The nucleotide sequence of the Klebsiella pneumoniae nifD gene is presented and together with the accompanying paper [Holland, Zilberstein, Zamir & Sussman (1987) Biochem. J. 247, 277-285] completes the sequence of the nifHDK genes encoding the nitrogenase polypeptides. The K. pneumoniae nifD gene encodes the 483-amino acid-residue nitrogenase alpha-subunit polypeptide of Mr 54156. The alpha-subunit has five strongly conserved cysteine residues at positions 63, 89, 155, 184 and 275, some occurring in a region showing both primary sequence and potential structural homology to the K. pneumoniae nitrogenase beta-subunit. A comparison with six other alpha-subunit amino acid sequences has been made, which indicates a number of potentially important domains within alpha-subunits.     

Nucleotide sequence of a chicken vitellogenin gene and derived amino acid sequence of the encoded yolk precursor protein

The gene encoding the major vitellogenin from chicken has been completely sequenced and its exon-intron organization has been established. The gene is 20,342 base-pairs long and contains 35 exons with a combined length of 5787 base-pairs. They encode the 1850-amino acid pre-peptide of vitellogenin, which is the precursor of the mature yolk proteins, the serine-rich and heavily phosphorylated phosvitin and the lipovitellin. The 217-amino acid phosvitin polypeptide occupies an internal position (residue 1112 through 1328) within the vitellogenin molecule. The 125,000 and 30,000 Mr lipovitellin polypeptides are encoded by the sequences at the N-terminal and the C-terminal sides of the phosvitin section, respectively. The main features of the gene and protein sequences, and the evolutionary implications, are discussed.     

Identification of amino acid substitutions that alter the substrate specificity of TEM-1 beta-lactamase.

TEM-1 beta-lactamase is the most prevalent plasmid-mediated beta-lactamase in gram-negative bacteria. Recently, TEM beta-lactamase variants with amino acid substitutions in the active-site pocket of the enzyme have been identified in natural isolates with increased resistance to extended-spectrum cephalosporins. To identify other amino acid substitutions that alter the activity of TEM-1 towards extended-spectrum cephalosporins, we probed regions around the active-site pocket by random-replacement mutagenesis. This mutagenesis technique involves randomizing the DNA sequence of three to six codons in the blaTEM-1 gene to form a library containing all or nearly all of the possible substitutions for the region randomized. In total, 20 different residue positions that had been randomized were screened for amino acid substitutions that increased enzyme activity towards the extended-spectrum cephalosporin cefotaxime. Substitutions at positions 104, 168, and 238 in the TEM-1 beta-lactamase that resulted in increased enzyme activity towards extended-spectrum cephalosporins were found. In addition, small deletions in the loop containing residues 166 to 170 drastically altered the substrate specificity of the enzyme by increasing activity towards extended-spectrum cephalosporins while virtually eliminating activity towards ampicillin.     

Relation between the structure of oligopeptides and the amino acid sequence

The present paper deals with determination of the relationship between the order of the arrangement of amino acids in comparatively short-range oligopeptides (tetrapeptides) and their conformational potentialities. It is shown that the spatial and conformational possibilities of the tetrapeptides composed of the same amino acid residues exhibit high sensibility to their mutual arrangement, i. e. to the amino acid sequence. A detailed conformation analysis vividly demonstrated that the difference in conformational possibilities is manly determined by different conditions of realization of residual interactions. It is shown convincingly that energetic differences of the fragments are due to different interaction contributions for each of the considered fragments.     

Complete amino acid sequence of human phosphoglycerate kinase. Isolation and amino acid sequence of tryptic peptides

As a part of the elucidation of the complete amino acid sequence of human phosphoglycerate kinase, 46 tryptic peptides, ranging in length from 1 to 26 residues, were isolated and characterized from the reduced and S-carboxymethylated enzyme. The isolated peptides were subjected to sequence analysis by the modified dansyl-Edman degradation procedure and automated Edman degradation technique. The results, together with the data on cyanogen bromide peptides and two additional tryptic peptides from cyanogen bromide peptides reported in the accompanying paper, established the complete amino acid sequence of human erythrocyte phosphoglycerate kinase.     

Saccharification and fermentation of Sugar Cane bagasse by Klebsiella oxytoca P2 containing chromosomally integrated genes encoding the Zymomonas mobilis ethanol pathway

Pretreatment of sugar cane bagasse is essential for a simultaneous saccharification and fermentation (SSF) process which uses recombinant Klebsiella oxytoca strain P2 and Genencor Spezyme CE. Strain P2 has been genetically engineered to express Zymomonas mobilis genes encoding the ethanol pathway and retains the native ability to transport and metabolize cellobiose (minimizing the need for extracellular cellobiase). In SSF studies with this organism, both the rate of ethanol production and ethanol yield were limited by saccharification at 10 and 20 filter papaer units (FPU) g(-1) acid-treated bagasse. Dilute slurries of biomass were converted to ethanol more efficiently (over 72% of theoretical yield) in simple batch fermentations than slurries containing high solids albeit with the production of lower levels of ethanol. With high solids (i.e., 160 g acid-treated bagasse L(-1)), a combination of 20 FPU cellulase g(-1) bagasse, preincubation under saccharification conditions, and additional grinding (to reduce particle size) were required to produce ca. 40 g ethanol L(-1). Alternatively, almost 40 g ethanol L(-1) was produced with 10 FPU cellulase g(-1) bagasse by incorporating a second saccharification step (no further enzyme addition) followed by a second inoculation and short fermentation. In this way, a theoretical ethanol yield of over 70% was achieved with the production of 20 g ethanol 800 FPU(-1) of commercial cellulase. (c) 1994 John Wiley & Sons, Inc.