An improved protocol for rapid extraction of membrane enzymes from Gram positive bacteria

Attri, Pooja; Jodha, Drukshakshi; Singh, Jasbir; Dhanda, Suman

doi:10.1039/c2ay05931b

Cited by 5 publications

(4 citation statements)

References 16 publications

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“…The results obtained by site-directed mutagenesis of the LsbB bacteriocin are consistent with data obtained by other laboratories, where most amino acid substitutions cause a reduction in or complete loss of activity. It is interesting, however, that some substitutions, such as alanine to serine (Ala 30 Ser), enhanced bacteriocin activity, most likely due to better interaction and positioning in the YvjB receptor. In addition, we suggest that not only residues Trp 25 and Ala 30 , but also the distance between them, play an essential role in the antimicrobial activity of LsbB.…”

Section: Discussionmentioning

confidence: 99%

“…First, using total DNA BGMN1-596 and designed primers ex1 SacI/ HindIII for YvjBex1, and ex2 BamHI/HindIII for YvjBex2, corresponding fragments were amplified by PCR. The PCR products were purified and cloned first to pGEM-T-Easy vector, confirmed by sequencing, and recloned as SacI/HindIII or BamHI/HindIII in frame with 6ϫHis tag to pQE 30 expression vector. Fusion His-tagged proteins were expressed in E. coli M15 cells.…”

Section: Methodsmentioning

confidence: 99%

“…Previous work indicates that the last residue, Ala 30 , is important for bacteriocin activity. As expected, the deletion of the terminal Ala 30 , the addition of another alanine at the end (Ala 31 ), or the substitution of Ala 30 with Pro caused a total loss of antimicrobial activity of LsbB, whereas substitutions of Ala 30 with other residues, such as Gly and Val, had no apparent impact on the bacteriocin activity, while substitution with Ser enhanced bacteriocin activity (Fig. 1A).…”

Section: Lsbb Causes Membrane Damage Of Sensitive Cellsmentioning

confidence: 99%

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LsbB Bacteriocin Interacts with the Third Transmembrane Domain of the YvjB Receptor

Miljković

Uzelac

Mirković

et al. 2016

Appl Environ Microbiol

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The Zn-dependent membrane-located protease YvjB has previously been shown to serve as a target receptor for LsbB, a class II leaderless lactococcal bacteriocin. Although yvjB is highly conserved in the genus Lactococcus, the bacteriocin appears to be active only against the subspecies L. lactis subsp. lactis. Comparative analysis of the YvjB proteins of a sensitive strain (YvjB MN ) and a resistant strain (YvjB MG ) showed that they differ from each other in 31 positions. In this study, we applied site-directed mutagenesis and performed directed binding studies to provide biochemical evidence that LsbB interacts with the third transmembrane helix of YvjB in susceptible cells. The site-directed mutagenesis of LsbB and YvjB proteins showed that certain amino acids and the length of LsbB are responsible for the bacteriocin activity, most probably through adequate interaction of these two proteins; the essential amino acids in LsbB responsible for the activity are tryptophan ( Bacteriocins are small, ribosomally synthesized, cationic, and hydrophobic peptides produced by various bacteria, and they are often found to be active against bacteria closely related to the producers. However, some also have broader inhibitory spectra, including pathogens and problematic bacteria. Producer organisms are immune to their own bacteriocin(s), a property that is mediated by specific immunity proteins (1).Bacteriocins from Gram-positive bacteria are generally classified into two main groups: the class I lantibiotics, containing posttranslationally modified peptides with ring-forming lanthionine or methyllanthionine residues, and class II, composed of nonmodified or minimally modified peptide bacteriocins (1-3). Class II bacteriocins are further subdivided into pediocin-like bacteriocins (class IIa), two-peptide bacteriocins (class IIb), circular bacteriocins (class IIc), and nonpediocin one-peptide bacteriocins (class IId) (2).Bacteriocins have been much studied from a fundamental and scientific perspective and also for their potential applications as food preservatives, and in veterinary and human medicine as alternatives to antibiotics or as synergists (1, 4).Bacteriocins have a number of positive attributes that have made them especially attractive for various applications (5). Some of them exhibit a broad spectrum of activity, inhibiting microorganisms belonging to different genera and species, including many bacterial pathogens that cause human, animal, or plant infections (6). For clinical applications, bacteriocins have been presented as a viable alternative to antibiotics due to the high specificity of certain bacteriocins against clinical pathogens, including multidrug-resistant (MDR) strains (4). Therefore, these substances have various potential applications in the food industry and medicine, either alone or in combination with other chemicals or methods (7).The multiplicity and diversity of bacteriocins and the resultant effects of their interactions with targeted bacteria on microbial ecology have been thoro...

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Lsbb Causes Membrane Damage Of Sensitive Cellsmentioning

confidence: 99%

See 1 more Smart Citation

LsbB Bacteriocin Interacts with the Third Transmembrane Domain of the YvjB Receptor

Miljković

Uzelac

Mirković

et al. 2016

Appl Environ Microbiol

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“…This will confirm whether the key anti-cancer component of IB is present in its outer membrane. Going further, the two most abundant groups of surface molecules found on C. sporogenes, proteoglycans [250] and cell wall proteins [251], The total levels and/or phosphorylation status of individual cyclins can be easily and rapidly measured using specific antibodies by immunoblotting. The Cyclin D1…”

Section: Cancer Activitymentioning

confidence: 99%

Bacteria-mediated anti-cancer therapy

Bhave¹

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Membrane Bound Aminopeptidase B of a Potential Probiotic Pediococcus acidilactici NCDC 252: Purification, Physicochemical and Kinetic Characterization

Attri

Jodha

Bansal

et al. 2021

Int J Pept Res Ther

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An improved protocol for rapid extraction of membrane enzymes from Gram positive bacteria

Cited by 5 publications

References 16 publications

LsbB Bacteriocin Interacts with the Third Transmembrane Domain of the YvjB Receptor

LsbB Bacteriocin Interacts with the Third Transmembrane Domain of the YvjB Receptor

Bacteria-mediated anti-cancer therapy

Membrane Bound Aminopeptidase B of a Potential Probiotic Pediococcus acidilactici NCDC 252: Purification, Physicochemical and Kinetic Characterization

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