Dataset Information

Identification of a PutP proline permease gene homolog from Staphylococcus aureus by expression cloning of the high-affinity proline transport system in Escherichia coli.

ABSTRACT: The important food-borne pathogen Staphylococcus aureus is distinguished by its ability to grow at low water activity values. Previous work in our laboratory and by others has revealed that proline accumulation via transport is an important osmoregulatory strategy employed by this bacterium. Furthermore, proline uptake by this bacterium has been shown to be mediated by two distinct transport systems: a high-affinity system and a low-affinity system (J.-H. Bae, and K. J. Miller, Appl. Environ. Microbiol. 58:471-475, 1992; D. E. Townsend and B. J. Wilkinson, J. Bacteriol. 174:2702-2710, 1992). In the present study, we report the cloning of the high-affinity proline transport system of S. aureus by functional expression in an Escherichia coli host. The sequence of the staphylococcal proline permease gene was predicted to encode a protein of 497 amino acids which shares 49% identity with the PutP high-affinity proline permease of E. coli. Analysis of hydropathy also indicated a common overall structure for these proteins.

SUBMITTER: Wengender PA

PROVIDER: S-EPMC167280 | biostudies-other | 1995 Jan

REPOSITORIES: biostudies-other

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Identification of a PutP proline permease gene homolog from Staphylococcus aureus by expression cloning of the high-affinity proline transport system in Escherichia coli.

Wengender P A PA Miller K J KJ

Applied and environmental microbiology 19950101 1

The important food-borne pathogen Staphylococcus aureus is distinguished by its ability to grow at low water activity values. Previous work in our laboratory and by others has revealed that proline accumulation via transport is an important osmoregulatory strategy employed by this bacterium. Furthermore, proline uptake by this bacterium has been shown to be mediated by two distinct transport systems: a high-affinity system and a low-affinity system (J.-H. Bae, and K. J. Miller, Appl. Environ. Mi ...[more]

PMID: 7887605

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Project description:Staphylococcus aureus causes a wide variety of invasive human infections. However, delineation of the genes which are essential for the in vivo survival of this pathogen has not been accomplished to date. Using signature tag mutagenesis techniques and large mutant pool screens, previous investigators identified several major gene classes as candidate essential gene loci for in vivo survival; these include genes for amino acid transporters, oligopeptide transporters, and lantibiotic synthesis (W. R. Schwan, S. N. Coulter, E. Y. W. Ng, M. H. Langhorne, H. D. Ritchie, L. L. Brody, S. Westbrock-Wadman, A. S. Bayer, K. R. Folger, and C. K. Stover, Infect. Immun. 66:567-572, 1998). In this study, we directly compared the virulence of four such isogenic signature tag mutants with that of the parental strain (RN6390) by using a prototypical model of invasive S. aureus infection, experimental endocarditis (IE). The oligonucleotide signature tag (OST) mutant with insertional inactivation of the gene (putP) which encodes the high-affinity transporter for proline uptake exhibited significantly reduced virulence in the IE model across three challenge inocula (10(4) to 10(6) CFU) in terms of achievable intravegetation densities (P, <0.05). The negative impact of putP inactivation on in vivo survival in the IE model was confirmed by simultaneous challenge with the original putP mutant and the parental strain as well as by challenge with a putP mutant in which this genetic inactivation was transduced into a distinct parental strain (S6C). In contrast, inactivation of loci encoding an oligopeptide transporter, a purine repressor, and lantibiotic biosynthesis had no substantial impact on the capacity of OST mutants to survive within IE vegetations. Thus, genes encoding the uptake of essential amino acids may well represent novel targets for new drug development. These data also confirm the utility of the OST technique as an important screening methodology for identifying candidate genes as requisite loci for the in vivo survival of S. aureus.

Project description:Escherichia coli and Staphylococcus aureus are important agents of urinary tract infections that can often evolve to severe infections. The rise of antibiotic-resistant strains has driven the search for novel therapies to replace the use or act as adjuvants of antibiotics. In this context, plant-derived compounds have been widely investigated. Cuminaldehyde is suggested as the major antimicrobial compound of the cumin seed essential oil. However, this effect is not fully understood. Herein, we investigated the in silico and in vitro activities of cuminaldehyde, as well as its ability to potentiate ciprofloxacin effects against S. aureus and E. coli. In silico analyses were performed by using different computational tools. The PASS online and SwissADME programmes were used for the prediction of biological activities and oral bioavailability of cuminaldehyde. For analysis of the possible toxic effects and the theoretical pharmacokinetic parameters of the compound, the Osiris, SwissADME and PROTOX programmes were used. Estimations of cuminaldehyde gastrointestinal absorption, blood brain barrier permeability and skin permeation by using SwissADME; and drug likeness and score by using Osiris, were also evaluated The in vitro antimicrobial effects of cuminaldehyde were determined by using microdilution, biofilm formation and time-kill assays. In silico analysis indicated that cuminaldehyde may act as an antimicrobial and as a membrane permeability enhancer. It was suggested to be highly absorbable by the gastrointestinal tract and likely to cross the blood brain barrier. Also, irritative and harmful effects were predicted for cuminaldehyde if swallowed at its LD50. Good oral bioavailability and drug score were also found for this compound. Cuminaldehyde presented antimicrobial and anti-biofilm effects against S. aureus and E. coli.. When co-incubated with ciprofloxacin, it enhanced the antibiotic antimicrobial and anti-biofilm actions. We suggest that cuminaldehyde may be useful as an adjuvant therapy to ciprofloxacin in S. aureus and E. coli-induced infections.

Dataset Information

Identification of a PutP proline permease gene homolog from Staphylococcus aureus by expression cloning of the high-affinity proline transport system in Escherichia coli.

Publications

Identification of a PutP proline permease gene homolog from Staphylococcus aureus by expression cloning of the high-affinity proline transport system in Escherichia coli.

Similar Datasets

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