scholarly journals Enterolysin A, a Cell Wall-Degrading Bacteriocin from Enterococcus faecalis LMG 2333

2003 ◽  
Vol 69 (5) ◽  
pp. 2975-2984 ◽  
Author(s):  
Trine Nilsen ◽  
Ingolf F. Nes ◽  
Helge Holo
Keyword(s):  
Amino Acids ◽  
Cell Wall ◽  
Amino Acid ◽  
Enterococcus Faecalis ◽  
Antimicrobial Protein ◽  
Terminal Part ◽  
Calculated Molecular Weight ◽  
Solid Media ◽  
Dependent Signal ◽  
Modular Structures

ABSTRACT A novel antimicrobial protein, designated enterolysin A, was purified from an Enterococcus faecalis LMG 2333 culture. Enterolysin A inhibits growth of selected enterococci, pediococci, lactococci, and lactobacilli. Antimicrobial activity was initially detected only on solid media, but by growing the bacteria in a fermentor under optimized production conditions (MRS broth with 4% [wt/vol] glucose, pH 6.5, and a temperature between 25 and 35°C), the bacteriocin activity was increased to 5,120 bacteriocin units ml−1. Enterolysin A production was regulated by pH, and activity was first detected in the transition between the logarithmic and stationary growth phases. Killing of sensitive bacteria by enterolysin A showed a dose-response behavior, and the bacteriocin has a bacteriolytic mode of action. Enterolysin A was purified, and the primary structure was determined by combined amino acid and DNA sequencing. This bacteriocin is translated as a 343-amino-acid preprotein with an sec-dependent signal peptide of 27 amino acids, which is followed by a sequence corresponding to the N-terminal part of the purified protein. Mature enterolysin A consists of 316 amino acids and has a calculated molecular weight of 34,501, and the theoretical pI is 9.24. The N terminus of enterolysin A is homologous to the catalytic domains of different cell wall-degrading proteins with modular structures. These include lysostaphin, ALE-1, zoocin A, and LytM, which are all endopeptidases belonging to the M37 protease family. The N-terminal part of enterolysin A is linked by a threonine-proline-rich region to a putative C-terminal recognition domain, which shows significant sequence identity to two bacteriophage lysins.

scholarly journals The structure of a glycopeptide isolated from the yeast cell wall

1968 ◽  
Vol 109 (3) ◽  
pp. 419-432 ◽  
Author(s):  
R. Sentandreu ◽  
D. H. Northcote
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Cell Wall ◽  
Amino Acid ◽  
Yeast Cell ◽  
Aspartic Acid ◽  
Yeast Cell Wall ◽  
Low Molecular Weight ◽  
Elimination Reaction ◽  
Sephadex Column

1. Glycopeptides containing mannose were extracted from isolated yeast cell walls by ethylenediamine and purified by treatment with Pronase and fractionation on a Sephadex column. 2. A glycopeptide that appeared homogeneous on electrophoresis and ultracentrifugation had a molecular weight of 76000, and contained a high-molecular-weight mannan and approx. 4% of amino acids. 3. The amino acid composition of the peptide was determined. It was rich in serine and threonine and also contained glucosamine. No cystine and methionine were detected. 4. The glycopeptide underwent a β-elimination reaction when treated with dilute alkali at low temperatures. The reaction resulted in the release of mannose, mannose disaccharides and possibly other low-molecular-weight mannose oligosaccharides. During the β-elimination reaction the dehydro derivatives of serine and threonine were formed. One of the linkages between carbohydrate and amino acids in the glycopeptide is an O-mannosyl bond from mannose and mannose oligosaccharides to serine and threonine. 5. After the β-elimination reaction the bulk of the mannose in the form of the large mannan component was still covalently linked to the peptide. This polysaccharide was therefore attached to the amino acids by a linkage different from the O-mannosyl bonds to serine and threonine that attach the low-molecular-weight sugars. 6. Mannan was prepared from the glycopeptide and from the yeast cell wall by treatment of the fractions with hot solutions of alkali. The mannan contained aspartic acid and glucosamine and some other amino acids. The aspartic acid and glucosamine were present in equimolar amounts; the aspartic acid was the only amino acid present in an amount equivalent to that of glucosamine. Thus there is the possibility of a linkage between the mannan and the peptide via glucosamine and aspartic acid. 7. Mannose 6-phosphate was shown to be part of the mannan structure. Information about the structure of the mannan and the linkage of the glucosamine was obtained by periodate oxidation studies. 8. The glucosamine present in the glycopeptide could not be released by treatment with an enzyme preparation obtained from the gut of Helix pomatia. This enzyme released glucosamine from the intact cell wall. Thus there are probably at least two polymers containing glucosamine in the cell wall. 9. The biosynthesis of the mannan polymer in the yeast cell wall is discussed with regard to the two types of carbohydrate–amino acid linkages found in the glycoprotein.

Simplified growth media for Entomophthora egressa protoplasts

1982 ◽  
Vol 28 (7) ◽  
pp. 815-821 ◽  
Author(s):  
Gary B. Dunphy ◽  
Richard A. Nolan
Keyword(s):  
Amino Acids ◽  
Cell Wall ◽  
Amino Acid ◽  
Fetal Calf Serum ◽  
Calf Serum ◽  
Normal Growth ◽  
Cysteic Acid ◽  
Growth Media ◽  
Cell Wall Regeneration ◽  
Glucose Medium

The protoplast stage of Entomophthora egressa utilized cysteic acid, L-isoleucine, L-leucine, L-methionine, L-proline, DL-serine, L-threonine, L-glutamine and (or) L-asparagine, and L-valine during the initial 16 h of incubation in modified Grace's medium but did not utilize glucose. Medium A, containing the above amino acids (excluding L-isoleucine) plus L-tyrosine, L-glutamic acid, L-aspartic acid, and L-lysine, supported normal growth and cell wall regeneration. The amino acid composition was further simplified to include only L-glutamine, L-asparagine, and L-methionine (medium B). The growth rates and morphogenesis of the protoplasts in media A and B were compared with those in medium A plus albumin. Fetal calf serum was essential for protoplast growth in shaken cultures in both simplified media; however, it was not required for growth in stationary cultures. Depending on the medium employed, E. egressa exhibited complete or partial vitamin auxoautotrophy.

scholarly journals Diversity of ace, a Gene Encoding a Microbial Surface Component Recognizing Adhesive Matrix Molecules, from Different Strains of Enterococcus faecalis and Evidence for Production of Ace during Human Infections

2000 ◽  
Vol 68 (9) ◽  
pp. 5210-5217 ◽  
Author(s):  
Sreedhar R. Nallapareddy ◽  
Kavindra V. Singh ◽  
Ruay-Wang Duh ◽  
George M. Weinstock ◽  
Barbara E. Murray
Keyword(s):  
Amino Acids ◽  
Extracellular Matrix ◽  
Amino Acid ◽  
Enterococcus Faecalis ◽  
Extracellular Matrix Proteins ◽  
Matrix Proteins ◽  
Rabbit Serum ◽  
Collagen Types ◽  
Collagen Binding ◽  
Different Strains

ABSTRACT Our previous work reported that most Enterococcus faecalis strains adhered to the extracellular matrix proteins collagen types I and IV and laminin after growth at 46°C, but not 37°C, and we subsequently identified an E. faecalissequence, ace, that encodes a bacterial adhesin similar to the collagen binding protein Cna of Staphylococcus aureus. In this study, we examined the diversity of E. faecalis-specific ace gene sequences among different isolates obtained from various geographic regions as well as from various clinical sources. A comparison of nucleotide and deduced amino acid sequences of Ace from nine E. faecalis strains identified a highly conserved N-terminal A domain, followed by a variable B domain which contains two to five repeats of 47 amino acids in tandem array, preceded by a 20-amino-acid partial repeat. Using 17 other strains collected worldwide, the 5′ region of acethat encodes the A domain was sequenced, and these sequences showed ≥97.5% identity. Among the previously reported five amino acids critical for collagen binding by Cna of S. aureus, four were found to be identical in Ace from all strains tested. Polyclonal immune rabbit serum prepared against recombinant Ace A derived fromE. faecalis strain OG1RF detected Ace in mutanolysin extracts of seven of nine E. faecalis strains after growth at 46°C; Ace was detected in four different molecular sizes that correspond to the variation in the B repeat region. To determine if there was any evidence to indicate that Ace might be produced under physiological conditions, we quantitatively assayed sera collected from patients with enterococcal infections for the presence of anti-Ace A antibodies. Ninety percent of sera (19 of 21) from patients withE. faecalis endocarditis showed reactivity with titers from 1:32 to >1:1,024; the only 2 sera which lacked antibodies to Ace A had considerably lower titers of antibodies to other E. faecalis antigens as well. Human-derived, anti-Ace A immunoglobulins G purified from an E. faecalis endocarditis patient serum inhibited adherence of 46°C-grown E. faecalis OG1RF to collagen types I and IV and laminin. In conclusion, these results show that ace is highly conserved among isolates of E. faecalis, with at least four variants related to the differences in the B domain, is expressed by different strains during infection in humans, and human-derived antibodies can block adherence to these extracellular matrix proteins.

scholarly journals Identification of Two Binding Domains, One for Peptidoglycan and Another for a Secondary Cell Wall Polymer, on the N-Terminal Part of the S-Layer Protein SbsB from Bacillus stearothermophilus PV72/p2

1998 ◽  
Vol 180 (24) ◽  
pp. 6780-6783 ◽  
Author(s):  
Margit Sára ◽  
Eva M. Egelseer ◽  
Christine Dekitsch ◽  
Uwe B. Sleytr
Keyword(s):  
Amino Acids ◽  
Cell Wall ◽  
Secondary Cell Wall ◽  
Bacillus Stearothermophilus ◽  
Binding Domain ◽  
Terminal Part ◽  
Binding Domains ◽  
Cell Wall Polymer ◽  
Function Relationship ◽  
Secondary Cell Wall Polymer

ABSTRACT First studies on the structure-function relationship of the S-layer protein from B. stearothermophilus PV72/p2 revealed the coexistence of two binding domains on its N-terminal part, one for peptidoglycan and another for a secondary cell wall polymer (SCWP). The peptidoglycan binding domain is located between amino acids 1 to 138 of the mature S-layer protein comprising a typical S-layer homologous domain. The SCWP binding domain lies between amino acids 240 to 331 and possesses a high serine plus glycine content.

Amino-acid-dependent signal transduction

2000 ◽  
Vol 351 (3) ◽  
pp. 545-550 ◽  
Author(s):  
Daphne A. VAN SLUIJTERS ◽  
Peter F. DUBBELHUIS ◽  
Edward F. C. BLOMMAART ◽  
Alfred J. MEIJER
Keyword(s):  
Signal Transduction ◽  
Amino Acids ◽  
Amino Acid ◽  
Metabolic Pathways ◽  
Short Review ◽  
Current State ◽  
Dependent Signal ◽  
Insulin Dependent

Recent research carried out in several laboratories has indicated that, in addition to their role as intermediates in many metabolic pathways, amino acids can interact with insulin-dependent signal transduction. In this short review, the current state of this rapidly expanding field is discussed.

scholarly journals Molecular Characterization of the S-Layer Gene, sbpA, of Bacillus sphaericus CCM 2177 and Production of a Functional S-Layer Fusion Protein with the Ability To Recrystallize in a Defined Orientation while Presenting the Fused Allergen

2002 ◽  
Vol 68 (7) ◽  
pp. 3251-3260 ◽  
Author(s):  
Nicola Ilk ◽  
Christine Völlenkle ◽  
Eva M. Egelseer ◽  
Andreas Breitwieser ◽  
Uwe B. Sleytr ◽  
...  
Keyword(s):  
Amino Acids ◽  
Cell Wall ◽  
Fusion Protein ◽  
Secondary Cell Wall ◽  
Bacillus Sphaericus ◽  
Terminal Part ◽  
Truncated Form ◽  
Sequence Encoding ◽  
Bet V1 ◽  
Secondary Cell Wall Polymer

ABSTRACT The nucleotide sequence encoding the crystalline bacterial cell surface (S-layer) protein SbpA of Bacillus sphaericus CCM 2177 was determined by a PCR-based technique using four overlapping fragments. The entire sbpA sequence indicated one open reading frame of 3,804 bp encoding a protein of 1,268 amino acids with a theoretical molecular mass of 132,062 Da and a calculated isoelectric point of 4.69. The N-terminal part of SbpA, which is involved in anchoring the S-layer subunits via a distinct type of secondary cell wall polymer to the rigid cell wall layer, comprises three S-layer-homologous motifs. For screening of amino acid positions located on the outer surface of the square S-layer lattice, the sequence encoding Strep-tag I, showing affinity to streptavidin, was linked to the 5′ end of the sequence encoding the recombinant S-layer protein (rSbpA) or a C-terminally truncated form (rSbpA31-1068). The deletion of 200 C-terminal amino acids did not interfere with the self-assembly properties of the S-layer protein but significantly increased the accessibility of Strep-tag I. Thus, the sequence encoding the major birch pollen allergen (Bet v1) was fused via a short linker to the sequence encoding the C-terminally truncated form rSpbA31-1068. Labeling of the square S-layer lattice formed by recrystallization of rSbpA31-1068/Bet v1 on peptidoglycan-containing sacculi with a Bet v1-specific monoclonal mouse antibody demonstrated the functionality of the fused protein sequence and its location on the outer surface of the S-layer lattice. The specific interactions between the N-terminal part of SbpA and the secondary cell wall polymer will be exploited for an oriented binding of the S-layer fusion protein on solid supports to generate regularly structured functional protein lattices.

scholarly journals Translation of Two Nested Genes in Bacteriophage P4 Controls Immunity-Specific Transcription Termination

1999 ◽  
Vol 181 (17) ◽  
pp. 5225-5233 ◽  
Author(s):  
Francesca Forti ◽  
Simona Polo ◽  
Kirk B. Lane ◽  
Erich W. Six ◽  
Gianpiero Sironi ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Nonsense Mutation ◽  
Transcription Termination ◽  
Open Reading Frame ◽  
Terminal Part ◽  
Reading Frame ◽  
Bacteriophage P4 ◽  
Nested Genes ◽  
Nonsense Codons

ABSTRACT In phage P4, transcription of the left operon may occur from both the constitutive PLE promoter and the regulated PLL promoter, about 400 nucleotides upstream of PLE. A strong Rho-dependent termination site,timm , is located downstream of both promoters. When P4 immunity is expressed, transcription starting at PLE is efficiently terminated attimm , whereas transcription from PLL is immunity insensitive and reads throughtimm . We report the identification of two nested genes, kil and eta, located in the P4 left operon. The P4 kil gene, which encodes a 65-amino-acid polypeptide, is the first translated gene downstream of the PLE promoter, and its expression is controlled by P4 immunity. Overexpression of kil causes cell killing. This gene is the terminal part of a longer open reading frame,eta, which begins upstream of PLE. Theeta gene is expressed when transcription starts from the PLL promoter. Three likely start codons predict a size between 197 and 199 amino acids for the Eta gene product. Bothkil and eta overlap thetimm site. By cloning kil upstream of a tRNA reporter gene, we demonstrated that translation of thekil region prevents premature transcription termination attimm . This suggests that P4 immunity might negatively control kil translation, thus enabling transcription termination at timm . Transcription starting from PLL proceeds throughtimm . Mutations that create nonsense codons ineta caused premature termination of transcription starting from PLL. Suppression of the nonsense mutation restored transcription readthrough at timm . Thus, termination of transcription from PLL is prevented by translation of eta.

scholarly journals ANTIBACTERIAL ACTIVITY ETHYL ACETATE EXTRACTS OF EARTHWORMS (Lumbricus rubellus, Eisenia foetida, Nereis sp) TOWARD Staphylococcus aureus, Enterococcus faecalis, Salmonella thyposa INVITRO

el–Hayah ◽  
2019 ◽  
Vol 7 (2) ◽  
pp. 62-73 ◽  
Author(s):  
Hartati Kartikaningsih ◽  
Sarastria Maharani ◽  
Fitarina Sartika
Keyword(s):  
Amino Acids ◽  
Staphylococcus Aureus ◽  
Antibacterial Activity ◽  
Amino Acid ◽  
Ethyl Acetate ◽  
Protein Content ◽  
Enterococcus Faecalis ◽  
Amino Acid Sequences ◽  
Lumbricus Rubellus ◽  
Eisenia Foetida

Earthworms had a mechanism of antibacterial. The research aimed to observe Lumbricus rubellus, Eisenia foetida, Nereis sp. antibacterial activity against Salmonella thyposa, Staphylococcus aureus, Enterococcus faecalis in vitro compared to ampicillin antibiotics. All the worms extracted using ethyl acetate extraction and tested their MIC. The compound of amino acids of the worms was analyzed by HPLC and nanodrop.  Lumbricus rubellus was the best anti-bacteria activity followed by Eisenia foetida and Nereis sp., but these activities less than ampicillin antibiotic. Observations with SEM showed these worms extract caused cell leakage in all of these bacteria. Protein content with Nanodrop testing revealed the highest protein content was Lumbricus rubellus (21.75 ppm) followed by Eisenia foetida (21.32 ppm) and Nereis sp. (20.98 ppm), as well as for amino acids levels, there were Lumbricus rubellus (24.66%), Eisenia foetida (22.78%), Nereis sp. (18.37%). From the 15 amino acids detected, all of the worms had the same sequence of fourth the highest amino acids (Glutamate, Aspartate, Leucine, Arginine) and fourth the lowest amino acid levels (Methionine, Hystidin, Tyrosin, Glisan). It had not been tested amino acid sequences of antibacterial compounds of these worms (Lumbricin 1: Phe-Ser-Lys-Tyr-Glu-Arg in Lumbricus rubellus worms, Fetidin 1: Ala-Met-Val-Ser-Ser and Fetidin 2: Ala-Met- Val-Gly-Thr in the Eisenia foetida worm, Hemerythrin: His-Glu-Asp in Nereis sp).

A Gastric Inhibitory Polypeptide II: The Complete Amino Acid Sequence

1971 ◽  
Vol 49 (8) ◽  
pp. 867-872 ◽  
Author(s):  
John C. Brown ◽  
Jill R. Dryburgh
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Amino Acid Residue ◽  
Gastric Inhibitory Polypeptide ◽  
Complete Amino Acid Sequence ◽  
Calculated Molecular Weight ◽  
Residue Polypeptide

Porcine gastric inhibitory polypeptide is a 43 amino acid residue polypeptide with the amino acid sequence Tyr–Ala–Glu–Gly–Thr–Phe–Ile–Ser–Asp–Tyr–Ser–Ile–Ala–Met–Asp–Lys–Ile–Arg–Gln–Gln–Asp–Phe–Val–Asn–Trp–Leu–Leu–Ala–Gln–Gln–Lys–Gly–Lys–Lys–Ser–Asp–Trp–Lys–His–Asn–Ile–Thr–Gln. Fifteen of the first 26 amino acids occur in the same position as they do in porcine glucagon, and nine of the first 26 in the same position as in porcine secretin. The calculated molecular weight of the polypeptide is 5105.

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