An Escherichia coli-Enterococcus faecalis shuttle vector as a tool for the construction of a group B Streptococcus heterologous mutant expressing the β antigen (Bac) of the C protein complex

Kreikemeyer, Bernd

doi:10.1016/s0378-1097(99)00496-6

Cited by 6 publications

(7 citation statements)

References 26 publications

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“…However, the ␤ protein encoded by one of these strains (24) is expressed at a very low level (31), which made it unclear whether size variation in the XPZ region is compatible with normal surface expression of the ␤ protein.…”

Section: Size Variation and Sequence Periodicity In The Xpz Regionmentioning

confidence: 99%

A Proline-Rich Region with a Highly Periodic Sequence in Streptococcal β Protein Adopts the Polyproline II Structure and Is Exposed on the Bacterial Surface

Areschoug

Linse²,

Stålhammar‐Carlemalm

et al. 2002

J Bacteriol

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Proline-rich regions have been identified in many surface proteins of pathogenic streptococci and staphylococci. These regions have been suggested to be located in cell wall-spanning domains and/or to be required for surface expression of the protein. Because little is known about these regions, which are found in extensively studied and biologically important surface proteins, we characterized the proline-rich region in one such protein, the ␤ protein of group B streptococci. The proline-rich region in ␤, designated the XPZ region, has a proline at every third position, and the sequence is highly periodic in other respects. Immunochemical analysis showed that the XPZ region was not associated with the cell wall but was exposed on the bacterial surface. Moreover, characterization of a ␤ mutant lacking the XPZ region demonstrated that this region was not required for surface expression of the ␤ protein. Comparison of the XPZ region in different ␤ proteins showed that it varied in size but always retained the typical sequence periodicity. Circular dichroism spectroscopy indicated that the XPZ region had the structure of a polyproline II helix, an extended and solvent-exposed structure with exactly three residues per turn. Because of the three-residue sequence periodicity in the XPZ region, it is expected to be amphipathic and to have distinct nonpolar and polar surfaces. This study identified a proline-rich structure with unique properties that is exposed on the surface of an important human pathogen.

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Section: Size Variation and Sequence Periodicity In The Xpz Regionmentioning

confidence: 99%

A Proline-Rich Region with a Highly Periodic Sequence in Streptococcal β Protein Adopts the Polyproline II Structure and Is Exposed on the Bacterial Surface

Areschoug

Linse²,

Stålhammar‐Carlemalm

et al. 2002

J Bacteriol

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“…Both proteins, which are mainly expressed by serotypes Ia, Ib and II, are thought to contribute significantly to virulence. The C protein b antigen (Cb protein) binds to the Fc portion of human IgA and may be important in bacterial resistance to mucosal immune defence mechanisms (Schalen, 1993;Kreikemeyer & Jerlstrom, 1999;Berner et al, 2002;Mawn et al, 1993). It is structurally related to pneumococcal protein Hic, which binds the complement inhibitor factor H and protects the organism from opsonophagocytosis (Jarva et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

Streptococcus agalactiae Cβ protein gene (bac) sequence types, based on the repeated region of the cell-wall-spanning domain: relationship to virulence and a proposed standardized nomenclature

Kong

Gidding

Berner

et al. 2006

Journal of Medical Microbiology

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The Cβ protein (Bac) of Streptococcus agalactiae (group B streptococcus; GBS) is an IgA binding protein encoded by bac, of which at least 39 sequence types have been described, based on polymorphisms in the repeated region of the cell-wall-spanning domain (‘bac sequence types’). Cβ is usually found in serotype Ib, less commonly in serotype II, and rarely in other serotypes. The aim of this study was to examine the prevalence, variety and distribution, among GBS serotypes and between invasive and superficial isolates, of bac sequence types. A total of 1101 GBS isolates were tested, from 10 countries, with a bac-specific PCR, and amplicons from all 255 (23 %) with positive results were sequenced. Ninety-seven percent (184/190) of serotype Ib and 37 % of serotype II isolates were bac positive. The Cα protein gene (bca) was present in 98 % (251/255), and insertion sequences IS1381 and IS861 in 94 % (239/255), of bac-positive isolates. The authors identified 59 bac sequence types belonging to 19 groups, based on length, from 496 to 946 bp, with up to six sequence variants (a–f) in each group. The median bac sequence length of invasive isolates was significantly shorter than that of superficial isolates overall (640 versus 586 bp; P <0.001) and specifically for serotype Ib (541 versus 676 bp; P <0.001), and invasive isolates were significantly (P <0.001) more likely to have one or more 18 bp deletions relative to the original published bac sequence (X59771). bac sequence typing is a useful addition to the previously described genotyping system, and will help to predict relative virulence among S. agalactiae serotype Ib strains.

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“…Consequently, the plasmid can be used for studies of enteric gram-negative bacteria, Streptococcus species (8,28), Enterococcus species (9), Streptococcus gordonii (unpublished results), L. lactis (10), Lactobacillus casei (12), and Pediococcus species (13). The copy number of the pAM401 parental plasmid (pGB354) in streptococci has been reported to be approximately 50 copies/cell (2).…”

Section: Resultsmentioning

confidence: 99%

Development of a Thermally Regulated Broad-Spectrum Promoter System for Use in Pathogenic Gram-Positive Species

Schofield

Westwater

Hoel

et al. 2003

Appl Environ Microbiol

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Selectively regulating gene expression is an essential molecular tool that is lacking for many pathogenic gram-positive bacteria. In this report, we describe the evaluation of a series of promoters regulated by the bacteriophage P1 temperature-sensitive C1 repressor in Enterococcus faecium, Enterococcus faecalis, and Staphylococcus aureus. Using the lacZ gene to monitor gene expression, we examined the strength, basal expression, and induced expression of synthetic promoters carrying C1 operator sites. The promoters exhibited extremely low basal expression and, under inducing conditions, gave high levels of expression (100-to 1,000-fold induction). We demonstrate that the promoter system could be modulated by temperature and showed rapid induction and that the mechanism of regulation occurred at the level of transcription. Controlled expression with the same constructs was also demonstrated in the gram-negative bacterium Escherichia coli. However, low basal expression and the ability to achieve derepression were dependent on both the number of mismatches in the C1 operator sites and the promoter driving c1 expression. Since the promoters were designed to contain conserved promoter elements from gram-positive species and were constructed in a broad-host-range plasmid, this system will provide a new opportunity for controlled gene expression in a variety of gram-positive bacteria.The overuse of antibiotics has contributed to the emergence and increasing prevalence of antibiotic-resistant bacteria. Gram-positive cocci such as Staphylococcus aureus and enterococci are the leading cause of hospital-acquired infections (39). S. aureus causes a variety of infections ranging from localized skin suppuration to life-threatening septicemia. Alarmingly, S. aureus isolates resistant to vancomycin, the last effective antibiotic, are emerging worldwide (22). Enterococcus species are a leading cause of urinary tract infection, nosocomial infection, and surgical-wound infection (39). Enterococcus faecalis is responsible for the majority of enterococcal infections (26,41) and, for the time being, usually remains sensitive to at least one antibiotic. In contrast Enterococcus faecium, which causes fewer infections, is more likely to be resistant to all antibiotics.The recent emergence of antibiotic-resistant gram-positive bacteria has highlighted the need for genetic studies addressing the mechanism of bacterial pathogenesis. Regulated promoters are essential for the functional analysis of genes through expression studies (1) and reverse genetics. However, only a few regulated promoters are available for use in enterococci and streptococci. The tetracycline-regulated promoter system has been shown to function in Streptococcus pneumoniae (47), Bacillus subtilis (11), and S. aureus (1, 25). In addition, the xylose-inducible promoter system has been used for B. subtilis and staphylococci (27,52,56). However, the levels of regulation achieved with these systems are below those obtained for gram-negative bacteria (33), and tight basal e...

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An Escherichia coli-Enterococcus faecalis shuttle vector as a tool for the construction of a group B Streptococcus heterologous mutant expressing the β antigen (Bac) of the C protein complex

Cited by 6 publications

References 26 publications

A Proline-Rich Region with a Highly Periodic Sequence in Streptococcal β Protein Adopts the Polyproline II Structure and Is Exposed on the Bacterial Surface

A Proline-Rich Region with a Highly Periodic Sequence in Streptococcal β Protein Adopts the Polyproline II Structure and Is Exposed on the Bacterial Surface

Streptococcus agalactiae Cβ protein gene (bac) sequence types, based on the repeated region of the cell-wall-spanning domain: relationship to virulence and a proposed standardized nomenclature

Development of a Thermally Regulated Broad-Spectrum Promoter System for Use in Pathogenic Gram-Positive Species

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