An Activity‐Based Probe for Studying Crosslinking in Live Bacteria

Gautam, Samir; Kim, Tae-Han; Shoda, Takuji; Sen, Sounok; Deep, Deeksha; Luthra, Ragini; Ferreira, Maria Teresa; Pinho, Mariana G.; Spiegel, David A.

doi:10.1002/anie.201503869

Cited by 34 publications

(31 citation statements)

References 31 publications

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“…50−52 Prior studies have demonstrated that structural mimicry of nascent PBP substrates results in PG incorporation in vitro (53) and in live bacterial cells. 54,55 Also, during the preparation of our manuscript it was shown, in vitro , that Ldts mediate cross-linking of synthetic Ldt substrates. 56 In contrast, there are currently no Ldt-specific probes to tag and visualize Ldt activity in live cells.…”

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confidence: 84%

L,D-Transpeptidase Specific Probe Reveals Spatial Activity of Peptidoglycan Cross-Linking

et al. 2019

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Peptidoglycan (PG) is a cross-linked, meshlike scaffold endowed with the strength to withstand the internal pressure of bacteria. Bacteria are known to heavily remodel their peptidoglycan stem peptides, yet little is known about the physiological impact of these chemical variations on peptidoglycan cross-linking. Furthermore, there are limited tools to study these structural variations, which can also have important implications on cell wall integrity and host immunity. Cross-linking of peptide chains within PG is an essential process, and its disruption thereof underpins the potency of several classes of antibiotics. Two primary cross-linking modes have been identified that are carried out by D,D-transpeptidases and L,D-transpeptidases (Ldts). The nascent PG from each enzymatic class is structurally unique, which results in different cross-linking configurations. Recent advances in PG cellular probes have been powerful in advancing the understanding of D,D-transpeptidation by Penicillin Binding Proteins (PBPs). In contrast, no cellular probes have been previously described to directly interrogate Ldt function in live cells. Herein, we describe a new class of Ldt-specific probes composed of structural analogs of nascent PG, which are metabolically incorporated into the PG scaffold by Ldts. With a panel of tetrapeptide PG stem mimics, we demonstrated that subtle modifications such as amidation of iso-Glu can control PG cross-linking. Ldt probes were applied to quantify and track the localization of Ldt activity in Enterococcus faecium, Mycobacterium smegmatis, and Mycobacterium tuberculosis. These results confirm that our Ldt probes are specific and suggest that the primary sequence of the stem peptide can control Ldt cross-linking levels. We anticipate that unraveling the interplay between Ldts and other cross-linking modalities may reveal the organization of the PG structure in relation to the spatial localization of cross-linking machineries.

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confidence: 84%

L,D-Transpeptidase Specific Probe Reveals Spatial Activity of Peptidoglycan Cross-Linking

et al. 2019

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“…The potency of the organic guanidine single crystals exposed to antibacterial drug was evaluated against two pathogenic bacterial strains. The biological activities of the Guanidine derivatives have been studied for its antibacterial activities using disk diffusion method 10 13 also confirmed the antibacterial activity of S. aerus against 2-Chloro-5-Ethoxy-3,6-Bis(Methylamino)-1,4-Benzoquinone single crystals. The activity of the Guanidine compounds towards Bacillus subtilis decreases in the order GuAce > GuAcr > Gutfb = Kanamycin > Guhtt= Gumal.…”

Section: Resultsmentioning

confidence: 74%

“…The antibacterial activity of the Guanidine derivatives.The Guanidine maleate and Guanidine tartarate shows a minimum activity against Bacillus subtilis bacteria and Guanidine tetrafluroborate shows the low activity against Staphylococcus aerus pathogen. The Guanidine derivatives with malaete and tartarate have less antibacterial activity against B. subtilis; and Guanidine derivatives with tetrafluroborate have less activity against S. aerus Gautam et al 2015. …”

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confidence: 98%

Antibacterial Activities of Guanidine Family Single Crystals against Bacillus Subtilis and Staphylococcus Aerus

2018

JCBSC

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A Guanidine derivative single crystal has been grown by simple ambient temperature gradient solution growth technique. The as grown single crystals tested against two bacterial pathogens like Bacillus subtilis, Staphylococcus aerus. The result shows that the single crystals have better resistance against the microbes. The standard antiseptic solution like kanamycin, it showed less or equal activity in pathogenic bacteria. In comparison to our currently used Guanidine derivatives single crystals, Guanidine Acetate shows better zone of inhibition against both the pathogens. But in contrast the Guanidine maleate and Guanidine Tetrafluroborate reveals lowest zone of inhibition against Bacillus subtilis and Staphylococcus aerus respectively. It can be concluded that Guanidine acetate exhibit excellent antibacterial effect and therefore, seems to be suitable as a local antiseptic agent, but further clinical studies are necessary.

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“…Scientists are still looking for probes targeting glycine cross-bridges (94), the membrane anchor of lipid II, or even the probes that can distinguish 3–3 and 3–4 cross-linking in PG. The search for novel PG probes will undoubtedly continue to be a very active area of research.…”

Section: Discussion and Future Directions In Visualization Of Peptidomentioning

confidence: 99%

Imaging Bacterial Cell Wall Biosynthesis

Radkov

Hsu

Booher

et al. 2018

Annu. Rev. Biochem.

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Peptidoglycan is an essential component of the cell wall that protects bacteria from environmental stress. A carefully coordinated biosynthesis of peptidoglycan during cell elongation and division is required for cell viability. This biosynthesis involves sophisticated enzyme machineries that dynamically synthesize, remodel, and degrade peptidoglycan. However, when and where bacteria build peptidoglycan, and how this is coordinated with cell growth, have been long-standing questions in the field. The improvement of microscopy techniques has provided powerful approaches to study peptidoglycan biosynthesis with high spatiotemporal resolution. Recent development of molecular probes further accelerated the growth of the field, which has advanced our knowledge of peptidoglycan biosynthesis dynamics and mechanisms. Here, we review the technologies for imaging the bacterial cell wall and its biosynthesis activity. We focus on the applications of fluorescent d-amino acids, a newly developed type of probe, to visualize and study peptidoglycan synthesis and dynamics, and we provide direction for prospective research.

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An Activity‐Based Probe for Studying Crosslinking in Live Bacteria

Cited by 34 publications

References 31 publications

L,D-Transpeptidase Specific Probe Reveals Spatial Activity of Peptidoglycan Cross-Linking

L,D-Transpeptidase Specific Probe Reveals Spatial Activity of Peptidoglycan Cross-Linking

Antibacterial Activities of Guanidine Family Single Crystals against Bacillus Subtilis and Staphylococcus Aerus

Imaging Bacterial Cell Wall Biosynthesis

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