A spore-lytic enzyme released from Bacillus cereus spores during germination

Makino, Shio; Ito, Norihito; Inoue, Toshihiko; Miyata, Seiko; Moriyama, Ryuichi

doi:10.1099/00221287-140-6-1403

Cited by 59 publications

(84 citation statements)

References 16 publications

(11 reference statements)

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“…The peptidoglycan binding motif of SleB is conserved many peptidoglycan hydrolases as mentioned above, while each hydrolase varies in the substrate specificity. The germination-related enzymes, such as SleB and SleC, recognize and hydrolyze only peptidoglycan with muramic δ-lactam moiety (Makino et al, 1994, Miyata et al, 1995a. The identification of the residues in the motif of the germination enzymes responsible for the recognition and/or binding to the muramic δ-lactam structure requires further study.…”

Section: Discussionmentioning

confidence: 99%

“…The muramic δ δ δ δ-lactam structure of cortex is crucial for the proper localization of SleB In our previous study, it was shown that SleB degrades only intact cortex but not the muramic δ-lactam-deficient cortex (Makino et al, 1994), suggesting that muramic δ-lactam is a major specific determinant of SleB as a germination lytic enzyme. Therefore, we assumed that muramic δ-lactam might play an important role in the localization process of SleB as well as in the substrate specificity of the enzyme.…”

Section: Construction Of Cwldgere Mutantmentioning

confidence: 99%

“…Such enzymes had been identified in the spores of Bacillus megaterium KM (GSLE) (Foster and Johnstone, 1987), Bacillus cereus IFO13597 (SleB) (Makino et al, 1994, Moriyama et al, 1996b, Bacillus subtilis 168 (SleB) (Moriyama et al, 1996a) and Clostridium perfringens S40 (SleC) (Miyata et al, 1995a, Miyata et al, 1995b. B. cereus SleB degrades only spore cortex peptidoglycan that contains muramic δ -lactam unique to the cortex, but not the peptidoglycan of vegetative cell walls (Makino et al, 1994, Miyata et al, 1995b. B. subtilis cwlD spores which completely lack muramic δ -lactam in their cortex are unable to complete germination through the blocking of cortex degradation (Sekiguchi et al, 1995, Popham et al, 1996.…”

Section: Introductionmentioning

confidence: 99%

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Subcellular Localization of a Germiantion-specific Cortex-lytic Enzyme, SleB, of Bacilli during Sporulation

et al. 2006

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The subcellular localization of a germination-specific cortex-lytic enzyme, SleB, of Bacillus subtilis during sporulation was observed by using fusions of N-terminal region of SleB to the green fluorescent protein (GFP). A fusion with a putative peptidoglycan-binding motif (SleB 1-108 -GFP) formed a fluorescent ring around the forespore of the wild type strain, as expected from the known location of the intact SleB in the dormant spore. SleB 1-108 -GFP formed a similar fluorescent ring around the forespore of the gerE mutant which has a severe defect in the coat structure, and of the cwlD mutant which lacks a muramic δ -lactam unique to the spore peptidoglycan (cortex), whereas the fusion could not attach to the spore of the cwlDgerE mutant. By contrast, a fusion without the motif (SleB 1-45 -GFP) could not be recruited around the forespore of the gerE mutant though it appeared to be accumulated on the outside of the spore of the wild type strain. Since SleB was shown to degrade only the cortex with muramic δ -lactam, these results suggested that a proper localization of SleB requires a strict interaction between the motif of the enzyme and the δ -lactam structure of the cortex, not the formation of normal coat layer.

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Section: Discussionmentioning

confidence: 99%

Section: Construction Of Cwldgere Mutantmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Subcellular Localization of a Germiantion-specific Cortex-lytic Enzyme, SleB, of Bacilli during Sporulation

et al. 2006

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“…Germination has been studied in an array of Gram-positive spore-forming species including Clostridium perfringens (Chen et al, 1997;Miyata et al, 1995), Bacillus thuringiensis (Hu et al, 2007), Bacillus subtilis (Ishikawa et al, 1998;Kodama et al, 1999;Moriyama et al, 1996a), Bacillus megaterium (Foster & Johnstone, 1987;Setlow et al, 2009), Bacillus cereus (Chen et al, 2000;Makino et al, 1994;Moriyama et al, 1996b) and B. anthracis (Heffron et al, 2009(Heffron et al, , 2010Lambert & Popham, 2008;Liu et al, 2004). The B. anthracis germination cascade begins when nutrient germinants contact receptors within the spore.…”

Section: Introductionmentioning

confidence: 99%

In vitro and in vivo analyses of the Bacillus anthracis spore cortex lytic protein SleL

2012

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The bacterial endospore is the most resilient biological structure known. Multiple protective integument layers shield the spore core and promote spore dehydration and dormancy. Dormancy is broken when a spore germinates and becomes a metabolically active vegetative cell. Germination requires the breakdown of a modified layer of peptidoglycan (PG) known as the spore cortex. This study reports in vitro and in vivo analyses of the Bacillus anthracis SleL protein.SleL is a spore cortex lytic enzyme composed of three conserved domains: two N-terminal LysM domains and a C-terminal glycosyl hydrolase family 18 domain. Derivatives of SleL containing both, one or no LysM domains were purified and characterized. SleL is incapable of digesting intact cortical PG of either decoated spores or purified spore sacculi. However, SleL derivatives can hydrolyse fragmented PG substrates containing muramic-d-lactam recognition determinants. The muropeptides that result from SleL hydrolysis are the products of N-acetylglucosaminidase activity. These muropeptide products are small and readily released from the cortex matrix. Loss of the LysM domain(s) decreases both PG binding and hydrolysis activity but these domains do not appear to determine specificity for muramic-d-lactam. When the SleL derivatives are expressed in vivo, those proteins lacking one or both LysM domains do not associate with the spore. Instead, these proteins remain in the mother cell and are apparently degraded. SleL with both LysM domains localizes to the coat or cortex of the endospore. The information revealed by elucidating the role of SleL and its domains in B. anthracis sporulation and germination is important in designing new spore decontamination methods. By exploiting germination-specific lytic enzymes, eradication techniques may be greatly simplified.

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“…The enzymic hydrolysis of spore-cortex peptidoglycan is essential for spores to complete rehydration during germination and to commence outgrowth. Spore cortex-lytic enzymes have been purified from spores of Bacillus megaterium (Foster & Johnstone, 1987), Bacillus cereus (Makino et al, 1994 ;Moriyama et al, 1996b ;Chen et al, 2000a, b) and Clostridium perfringens (Miyata et al, 1995), and were reviewed by . Based on structural analysis of spore peptidoglycan and its dynamics during germination, Atrih et al (1998 have suggested that there are at least three different types of enzyme activities involved in spore-cortex hydrolysis and modification during germination : glucosaminidase, lytic transglycosylase, and a possible non-hydrolytic epimerase.…”

Section: Introductionmentioning

confidence: 99%

Analysis of spore cortex lytic enzymes and related proteins in Bacillus subtilis endospore germination

et al. 2002

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The location and function of recognized cortex-lytic enzymes of Bacillus subtilis have been explored, and the involvement in germination of a number of related proteins tested. The SleB and CwlJ proteins are cortex-lytic enzymes, partially redundant in function, that are required together for effective cortex hydrolysis during B. subtilis spore germination. Spores were fractionated, and Western blotting of individual fractions suggests that the CwlJ protein is localized exclusively to the outer layers, or integument. The second spore-lytic enzyme, SleB, is localized both in the inner membrane of the spore and in the integument fraction. Neither protein changes location or size as the spore germinates. The ypeB gene is the second gene in a bicistronic operon with sleB. The SleB protein is absent from ypeB mutant spores, suggesting that YpeB is required for its localization or stabilization. In fractions of wild-type spores, the YpeB protein is found in the same locations as SleB -in both the inner membrane and the integument. As the absence of CwlJ protein does not affect the overall RP-HPLC profile of peptidoglycan fragments in germinating spores, this enzyme's hydrolytic specificity could not be defined. The effects of inactivation of several homologues of cortex-lytic enzymes of as yet undefined function were examined, by testing null mutants for their germination behaviour by OD 600 fall and by RP-HPLC of peptidoglycan fragments from dormant and germinating spores. The YaaH enzyme is responsible for a likely epimerase modification of peptidoglycan during spore germination, but the loss of this activity does not appear to affect the spore's ability to complete germination. Unlike the other cortex-lytic enzymes, the YaaH protein is present in large amounts in the spore germination exudate of B. subtilis. Mutants lacking either YdhD or YvbX, both homologues of YaaH, had no detectable alteration in either dormant or germinating spore peptidoglycan, and germinated normally. The ykvT gene, which encodes a protein of the SleB/CwlJ family, has no apparent association with germination : the gene is expressed in vegetative cells, and mutants lacking YkvT have no detectable phenotype.

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A spore-lytic enzyme released from Bacillus cereus spores during germination

Cited by 59 publications

References 16 publications

Subcellular Localization of a Germiantion-specific Cortex-lytic Enzyme, SleB, of Bacilli during Sporulation

Subcellular Localization of a Germiantion-specific Cortex-lytic Enzyme, SleB, of Bacilli during Sporulation

In vitro and in vivo analyses of the Bacillus anthracis spore cortex lytic protein SleL

Analysis of spore cortex lytic enzymes and related proteins in Bacillus subtilis endospore germination

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