Expression, secretion and antigenic variation of bacterial S-layer proteins.

The function of the S-layer, a regularly arranged structure on the outside of numerous bacteria, appears to be different for bacteria living in different environments. Almost no similarity exists between the primary sequences of S-proteins, although their amino acid composition is comparable. S-protein production is directed by single or multiple promoters in fron of the S-protein gene, yielding stable mRNAs. Most bacteria secrete S-proteins via the general secretory pathway (GSP). Translocation of S-protein across the outer membrane of Gram-negative bacteria sometimes occurs by S-protein-specific branches of the GSP. O-polysaccharide side-chains of the lipopolysaccharide component of the cell wall of Gram-negative bacteria appear to function as receptors for attachment of the S-layer. Silent S-protein genes have been found in Campylobacter fetus and Lactobaccillus acidophilus. These silent genes are placed in the expression site in a fraction of the bacterial population via inversiion of a chromosomal segment

A transcriptional barrier in the regulatory region of the tryptophan operon of Escherichia coli: its role in the regulation of repressor independent RNA synthesis

Chemicals/CAS: tryptophan, 6912-86-3, 73-22-3; RNA, Bacterial; RNA, Viral; Tryptophan, 73-22-3; Tryptophan-tRNA Ligase, EC 6.1.1.

Escherichia coli mutant strain with altered expression of the tryptophan operon: Ribonucleic acid synthesis in vitro

Chemicals/CAS: tryptophan, 6912-86-3, 73-22-3; Antibiotics, Aminoglycoside; DNA-Directed RNA Polymerases, EC 2.7.7.6; Heparin, 9005-49-6; Rifampin, 13292-46-1; RNA, Bacterial; Tryptophan, 73-22-

Avaliação funcional dos pacientes submetidos ao desbridamento artroscópico para tratamento das rupturas extensas e irreparáveis do manguito rotador Functional evaluation of patients who have undergone arthroscopic debridement to treat massive and irreparable tears of the rotator cuff

OBJETIVO: Avaliar os resultados dos pacientes submetidos ao desbridamento artroscópico das lesões extensas e irreparáveis do manguito rotador. Métodos: Foram operados 27 pacientes no período de 2003 a 2007, sendo avaliados 22 desses. O procedimento cirúrgico consistiu de desbridamento artroscópico do coto dos tendões envolvidos, bursectomia, remoção do osteófito acromial e, eventualmente, tenotomia do bíceps e tuberoplastia. RESULTADOS: No pré-operatório todos apresentavam envolvimento dos tendões do supra e infraespinal. Na avaliação pós-operatória, 14 pacientes estavam com o redondo menor íntegro e três com ruptura parcial do subescapular. Houve melhora dos critérios da UCLA de 15 no pré-operatório para 31 no pós. Não houve melhora de força muscular, porém ocorreu redução da dor. Conclusão: Desbridamento artroscópico é um procedimento indicado para pacientes idosos com ruptura irreparável do manguito rotador, que tenham boa ADM, baixa demanda funcional e com o principal objetivo de reduzir a dor.<br>OBJECTIVE: To evaluate the results in patients who have undergone arthroscopic debridement of massive and irreparable injury of the rotator cuff. METHODS: 27 patients were operated in the period from 2003 to 2007, during which 22 of them were evaluated. The procedure used consisted of arthroscopic debridement of the related tendons of the residual limb, bursectomy, acromial osteophyte removal, and eventually, biceps tenotomy and tuberoplasty. RESULTS: All patients showed involvement of the supraspinatus and infraspinatus tendons in the preoperative stage. In the postoperative evaluation, 14 patients had an teres minor muscle, and 3 had partial tears of the subscapularis tendon. There was an improvement in the UCLA criteria from 15 preoperatively to 31 postoperatively. There was no improvement in muscular strength, but there was a reduction in the pain. CONCLUSION: Arthroscopic debridement is a recommended procedure for elderly people with irreparable rotator cuff rupture, good ROM, low functional demand, and whose main objective is to diminish pain

Expression of Bacillus subtilis levanase gene in Lactobacillus plantarum and Lactobacillus casei

Two Lactobacillus-Escherichia coli shuttle vectors, harbouring the levanase gene from Bacillus subtilis under the control of its own promoter (pLPEW1) or behind the E. coli tac promoter (pE-SIEW2), were constructed. Lactobacillus plantarum showed the same growth characteristics on selective plates and in liquid media containing inulin, after transformation with either pLPEW1 or pESIEW2. L. plantarum transformed with pLPEW1 could be selected on inulin plates, indicating that levanase expression can be used as a food-grade selection system for Lactobacillus. Lactobacillus casei grew faster in inulin-containing medium than L. plantarum after transformation with pE-SIEW2, but did not grow when harbouring pLPEW1. Inulin-degrading activities of 90 mU/ml were found in culture medium of L. plantarum containing pLPEW1 or pESIEW2, and of 500 mU/ml in medium of L. casei (pESIEW2). Addition of 1 mM isopropyl β-D-thiogalactoside to the culture medium had no effect on growth and levanase expression in L. plantarum (pESIEW2) and L. casei (pESIEW2) strains. Levanase produced by L. casei (pESIEW2) has a size of 75 kDa and 72 kDa, corresponding to that of unprocessed and mature B. subtilis levanase, respectively, suggesting that the protein produced is recognized and processed by a signal peptidase

A convenient and reproducible method to genetically transform bacteria of the genus Bifidobacterium

A protocol was developed for the introduction of foreign plasmid DNA into various Bifidobacterium strains. The method, which is applicable to all Bifidobacterium species tested so far, is based on electroporation of bacteria made competent by preincubation in electroporation buffer for several hours at 4°C. Transformation of Bifidobacterium could be achieved with a plasmid vector originating from Bifidobacterium and with plasmid vectors from Corynebacterium, but not with vectors carrying replicons from Lactococcus or Lactobacillus

Structural and functional analysis of the S-layer protein crystallisation domain of Lactobacillus acidophilus ATCC 4356 : evidence for protein : protein interaction of two subdomains

The structure of the crystallisation domain, SAN, of the S A-protein of Lactobacillus acidophilus ATCC 4356 was analysed by insertion and deletion mutagenesis, and by proteolytic treatment. Mutant S A-protein synthesised in Escherichia coli with 7-13 amino acid insertions near the N terminus or within regions of sequence variation in SAN (amino acid position 7, 45, 114, 125, 193), or in the cell wall-binding domain (position 345) could form crystalline sheets, whereas insertions in conserved regions or in regions with predicted secondary structure elements (positions 30, 67, 88 and 156) destroyed this capacity. FACscan analysis of L. acidophilus synthesising three crystallising and one non-crystallising SA-protein c-myc (19 amino acid residues) insertion mutant was performed with c-myc antibodies. Fluorescence was most pronounced for insertions at positions 125 and 156, less for position 45 and severely reduced for position 7. By cytometric flow sorting a transformant harbouring the mutant SA-protein gene (position 125) was isolated that showed an increased fluorescense signal. Immunofluorescence microscopy suggested that the transformant synthesized mutant SA-protein only. PCR analysis of the transformant grown in the absence of selection pressure indicated that the mutant allele was stably integrated in the chromosome. Proteolytic treatment of SA-protein indicated that only sites near the middle of SAN are susceptible, although potential cleavage sites are present through the entire molecule. Expression in E. coli of DNA sequences encoding the two halves of SAN yielded peptides that could oligomerize. Our results indicate that SAN consists of a ∼12 kDa N and a ∼ 18 kDa C-terminal subdomain linked by a surface exposed loop. The capacity of SA-protein of L. acidophilus to present epitopes, up to ∼ 19 amino acid residues in length, at the bacterial surface in a genetically stable form, makes the system, in principle, suitable for application as an oral delivery vehicle. © 2002 Elsevier Science Ltd. All rights reserved. Chemicals/CAS: amino acid, 65072-01-7; DNA, 9007-49-2; Bacterial Proteins; Epitopes; Membrane Glycoproteins; Membrane Proteins; Peptide Fragments; Proto-Oncogene Proteins c-myc; Recombinant Proteins; surface array protein, bacteria; surface layer protein A, Bacteria; Vaccine