title: Activity and regulation of various forms of CwlJ, SleB, and YpeB proteins in degrading cortex peptidoglycan of spores of Bacillus species in vitro and during spore germination.
abstract: Germination of Bacillus spores requires degradation of a modified layer of peptidoglycan (PG) termed the spore cortex by two redundant cortex-lytic enzymes (CLEs), CwlJ and SleB, plus SleB's partner protein, YpeB. In this study, in vitro and in vivo analyses have been used to clarify the roles of individual SleB and YpeB domains in PG degradation. Purified mature Bacillus cereus SleB without its signal sequence (SleB(M)) and the SleB C-terminal catalytic domain (SleB(C)) efficiently triggered germination of decoated Bacillus megaterium and Bacillus subtilis spores lacking endogenous CLEs; previously, SleB's N-terminal domain (SleB(N)) was shown to bind PG but have no enzymatic activity. YpeB lacking its putative membrane anchoring sequence (YpeB(M)) or its N- and C-terminal domains (YpeB(N) and YpeB(C)) alone did not exhibit degradative activity, but YpeB(N) inhibited SleB(M) and SleB(C) activity in vitro. The severe germination defect of B. subtilis cwlJ sleB or cwlJ sleB ypeB spores was complemented by ectopic expression of full-length sleB [sleB(FL)] and ypeB [ypeB(FL)], but normal levels of SleB(FL) in spores required normal spore levels of YpeB(FL) and vice versa. sleB(FL) or ypeB(FL) alone, sleB(FL) plus ypeB(C) or ypeB(N), and sleB(C) or sleB(N) plus ypeB(FL) did not complement the cortex degradation defect in cwlJ sleB ypeB spores. In addition, ectopic expression of sleB(FL) or cwlJ(FL) with a Glu-to-Gln mutation in a predicted active-site residue failed to restore the germination of cwlJ sleB spores, supporting the role of this invariant glutamate as the key catalytic residue in SleB and CwlJ.
authors.affiliation: Department of Molecular, Microbial and Structural Biology, University of Connecticut Health Center, Farmington, Connecticut, USA.
authors.forename: Yunfeng
authors.initials: Y
authors.lastname: Li
authors.forename: Xuan Y
authors.initials: XY
authors.lastname: Butzin
authors.forename: Andrew
authors.initials: A
authors.lastname: Davis
authors.forename: Barbara
authors.initials: B
authors.lastname: Setlow
authors.forename: George
authors.initials: G
authors.lastname: Korza
authors.forename: Fatma Işik
authors.initials: FI
authors.lastname: Üstok
authors.forename: Graham
authors.initials: G
authors.lastname: Christie
authors.forename: Peter
authors.initials: P
authors.lastname: Setlow
authors.forename: Bing
authors.initials: B
authors.lastname: Hao
mesh_terms: D000581:Amidohydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D019943:Amino Acid Substitution; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D006867:Hydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007527:Isoenzymes; D050296:Microbial Viability; D009154:Mutation; D010457:Peptidoglycan / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D013171:Spores, Bacterial
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D007527:Isoenzymes; D010457:Peptidoglycan; C101656:SleB protein, Bacillus; D006867:Hydrolases; D000581:Amidohydrolases
references.citation: Microbiology (Reading). 2012 May;158(Pt 5):1359-1368
references.pmid: 22343356
references.citation: J Bacteriol. 1998 Mar;180(6):1375-80
references.pmid: 9515903
references.citation: Appl Environ Microbiol. 1989 Dec;55(12):3119-23
references.pmid: 16348073
references.citation: J Bacteriol. 2010 Oct;192(20):5378-89
references.pmid: 20729357
references.citation: J Bacteriol. 2011 Aug;193(16):4143-52
references.pmid: 21685283
references.citation: Proc Natl Acad Sci U S A. 1996 Dec 24;93(26):15405-10
references.pmid: 8986824
references.citation: Appl Microbiol Biotechnol. 1995 Mar;42(6):871-7
references.pmid: 7766087
references.citation: Microbiology (Reading). 2000 Jan;146 ( Pt 1):57-64
references.pmid: 10658652
references.citation: J Bacteriol. 2001 Aug;183(16):4886-93
references.pmid: 11466292
references.citation: Microbiology (Reading). 2001 Nov;147(Pt 11):2925-32
references.pmid: 11700343
references.citation: Microbiology (Reading). 2002 Aug;148(Pt 8):2383-2392
references.pmid: 12177332
references.citation: J Bacteriol. 2009 Sep;191(18):5569-76
references.pmid: 19581364
references.citation: Trends Microbiol. 2011 Feb;19(2):85-94
references.pmid: 21112786
references.citation: J Bacteriol. 1973 Nov;116(2):797-802
references.pmid: 4200857
references.citation: J Bacteriol. 2010 Jul;192(13):3424-33
references.pmid: 20435722
references.citation: J Bacteriol. 1996 Oct;178(20):6059-63
references.pmid: 8830707
references.citation: J Bacteriol. 2010 Feb;192(3):763-70
references.pmid: 19966006
references.citation: Proteins. 2012 Oct;80(10):2469-75
references.pmid: 22777830
references.citation: Trends Biochem Sci. 2004 Apr;29(4):169-72
references.pmid: 15124630
references.citation: Anal Chem. 2009 May 15;81(10):4035-42
references.pmid: 19374431
references.citation: Appl Environ Microbiol. 2012 Apr;78(8):2689-97
references.pmid: 22327596
references.citation: Nat Protoc. 2007;2(4):924-32
references.pmid: 17446874
references.citation: J Bacteriol. 2000 Mar;182(6):1499-506
references.pmid: 10692353
references.citation: Curr Opin Microbiol. 2003 Dec;6(6):550-6
references.pmid: 14662349
references.citation: Cell Mol Life Sci. 2002 Mar;59(3):426-33
references.pmid: 11964121
references.citation: J Bacteriol. 2001 Aug;183(16):4894-9
references.pmid: 11466293
references.citation: J Bacteriol. 2000 May;182(9):2513-9
references.pmid: 10762253
references.citation: J Bacteriol. 2009 Apr;191(7):2237-47
references.pmid: 19181808
references.citation: J Bacteriol. 2011 Jan;193(1):125-31
references.pmid: 20971910
references.citation: J Bacteriol. 2008 Dec;190(23):7601-7
references.pmid: 18835992
references.citation: J Bacteriol. 1996 Jun;178(12):3486-95
references.pmid: 8655545
references.citation: J Appl Microbiol. 2006 Sep;101(3):514-25
references.pmid: 16907802
references.citation: J Bacteriol. 2012 Sep;194(17):4537-45
references.pmid: 22730118
references.citation: J Bacteriol. 2000 Oct;182(19):5505-12
references.pmid: 10986255
references.citation: J Bacteriol. 2003 Apr;185(7):2315-29
references.pmid: 12644503
references.citation: Nat Methods. 2007 Sep;4(9):705-7
references.pmid: 17643108
references.citation: Genes Genet Syst. 2006 Jun;81(3):163-9
references.pmid: 16905870
references.citation: J Appl Microbiol. 2009 Jul;107(1):318-28
references.pmid: 19302310
references.citation: J Bacteriol. 2010 Sep;192(17):4317-26
references.pmid: 20581204