enzymology; Low molecular mass PBP; Bacillus subtilis
Abstract :
[en] Bacterial DD-peptidases are the targets of the β-lactam antibiotics. The sharp increase in bacterial resistance toward these antibiotics in recent years has stimulated the search for non-
β-lactam alternatives. The substrates of DD-peptidases are elements of peptidoglycan from bacterial cell walls. Attempts to base DD-peptidase inhibitor design on peptidoglycan structure, however, have not been particularly successful to date because the specific substrates for most of these enzymes are unknown. It is known, however, that the preferred substrates of low-molecular mass (LMM) class B and C DD-peptidases contain the free N-terminus of the relevant peptidoglycan. Two very similar LMMC enzymes, for example, the Actinomadura R39
DD-peptidase and Bacillus subtilis PBP4a, recognize a D-α-aminopimelyl terminus. The peptidoglycan of B. subtilis in the vegetative stage, however, has the N-terminal D-α-aminopimelyl carboxylic acid amidated. The question is, therefore, whether the DD-peptidases of B. subtilis are separately specific to carboxylate or carboxamide or have dual specificity. This paper describes an investigation of this issue with B. subtilis PBP4a. This enzyme was indeed found to have a dual specificity for peptide substrates, both in the acyl donor and in the acyl acceptor sites. In contrast, the R39 DD-peptidase, from an organism in which the peptidoglycan is not amidated, has a strong preference for a terminal carboxylate. It was also found that acyl acceptors, reacting with acyl−enzyme intermediates, were preferentially D-amino acid amides for PBP4a and the corresponding amino acids for the R39 DD-peptidase. Examination of the relevant crystal structures, aided by molecular modeling, suggested that the expansion of specificity in PBP4a accompanies a change of Arg351 in the R39 enzyme and most LMMC DD-peptidases to histidine in PBP4a and its orthologs in other Bacillus sp. This histidine, in neutral form at pH 7, appeared to be able to favorably interact with both carboxylate and carboxamide termini of substrates, in agreement with the kinetic data. It may still be possible, in specific cases, to combat bacteria with new antibiotics based on particular elements of their peptidoglycan structure.
Disciplines :
Microbiology
Author, co-author :
Nemmara, Venkatesh V.; Wesleyan university, connecticut 06459, USA > Department of chemistry
Adediran, S. A.; Wesleyan university, Connecticut 06459, USA > Department of chemistry
Dave, Kinjal; Wesleyan university, Connecticut 06459, USA > department of chemistry
Duez, Colette ; Université de Liège > Centre d'ingénierie des protéines
Pratt, Rex F.; Wesleyan university, Connecticut 06459, USA > Department of chemistry
Language :
English
Title :
Dual substrate specificity of Bacillus subtilis PBP4a
Publication date :
05 April 2013
Journal title :
Biochemistry
ISSN :
0006-2960
eISSN :
1520-4995
Publisher :
American Chemical Society, Washington, United States - District of Columbia
Waxman, D. J. and Strominger, J. L. (1983) Penicillin-binding proteins and the mechanism of action of β-lactam antibiotics Annu. Rev. Biochem. 52, 825-869
Macheboeuf, P., Contreras-Martel, C., Job, V., Dideberg, O., and Dessen, A. (2006) Penicillin-binding proteins: Key players in bacterial cell cycle and drug resistance processes FEMS Microbiol. Rev. 30, 673-691
Pratt, R. F. (2008) Substrate specificity of bacterial dd -peptidases (penicillin-binding proteins) Cell. Mol. Life Sci. 65, 2138-2155
Nemmara, V. V., Dzhekieva, L., Sarkar, K. S., Adediran, S. A., Duez, C., Nicholas, R. A., and Pratt, R. F. (2011) Substrate specificity of low-molecular mass bacterial dd -peptidases Biochemistry 50, 10091-10101
Popham, D. L. and Young, K. D. (2003) Role of penicillin-binding proteins in bacterial cell morphogenesis Curr. Opin. Microbiol. 6, 594-599
Matthei, P.-J., Neves, D., and Dessen, A. (2010) Bridging cell wall biosynthesis and bacterial morphogenesis Curr. Opin. Struct. Biol. 20, 749-755
Leyh-Bouille, M., Nakel, M., FreÌre, J.-M., Johnson, K., Duez, C., Ghuysen, J.-M., Nieto, M., and Perkins, H. R. (1972) Penicillin-sensitive dd -carboxypeptidases from Streptomyces strains R39 and K11 Biochemistry 11, 1290-1298
Ghuysen, J.-M., Leyh-Bouille, M., Campbell, J. N., Moreno, R., FreÌre, J.-M., Duez, C., Nieto, M., and Perkins, H. R. (1973) Structure of the wall peptidoglycan of Streptomyces R39 and the specificity and profile of its exocellular dd -carboxypeptidase-transpeptidase for peptide acceptors Biochemistry 12, 1243-1251
McDonough, M. A., Anderson, J. W., Silvaggi, N. R., Pratt, R. F., and Kelly, J. A. (2002) Structures of two kinetic intermediates reveal species specificity of penicillin-binding proteins J. Mol. Biol. 322, 111-122
Sauvage, E., Powell, A. J., Heilemann, J., Josephine, H. R., Charlier, P., Davies, C., and Pratt, R. F. (2008) Crystal structures of complexes of bacterial dd -peptidases with peptidoglycan-mimetic ligands; the substrate specificity puzzle J. Mol. Biol. 381, 383-393
Dzhekieva, L., Rocaboy, M., Kerff, F., Charlier, P., Sauvage, E., and Pratt, R. F. (2010) Crystal structure of a complex between the Actinomadura R39 dd -peptidase and a peptidoglycan-mimetic boronate inhibitor: Interpretation of a transition state analogue in terms of catalytic mechanism Biochemistry 49, 6411-6419
Sauvage, E., Duez, C., Herman, R., Kerff, F., Petrella, S., Anderson, J. W., Adediran, S. A., Pratt, R. F., FreÌre, J.-M., and Charlier, P. (2007) Crystal structure of the Bacillus subtilis penicillin-binding protein 4a and its complex with a peptidoglycan-mimetic peptide J. Mol. Biol. 371, 528-539
Atrih, A., Bacher, G., Allmaier, G., Williamson, M. P., and Foster, J. J. (1999) Analysis of peptidoglycan structure from vegetative cells of B. subtilis 168 and role of PBPs in peptidoglycan maturation J. Bacteriol. 181, 3956-3966
Warth, A. D. and Strominger, J. L. (1971) Structure of the peptidoglycan from vegetative cell walls of Bacillus subtilis J. Bacteriol. 180, 4967-4973
Warth, A. D. and Strominger, J. L. (1969) Structure of the peptidoglycan in bacterial spores: Occurrences of the lactam of muramic acid Proc. Natl. Acad. Sci. U.S.A. 64, 528-535
Bernard, E., Rolain, T., Courtin, P., Hols, P., and Chapot-Cartier, M.-P. (2011) Identification of the amidotransferase AsnB1 as being reasonable for meso-diaminopimelic acid amidation in Lactobacillus plantarum peptidoglycan J. Bacteriol. 193, 6323-6330
Pederson, L. B., Murray, T., Popham, D. L., and Setlow, P. (1998) Characterization of dacC, which encodes a new low-molecular weight penicillin-binding protein in Bacillus subtilis J. Bacteriol. 180, 4967-4973
Scheffers, D.-J. (2005) Dynamic localization of penicillin-binding proteins during spore development in Bacillus subtilis Microbiology 151, 999-1012
Anderson, J. W., Adediran, S. A., Charlier, P., Nguyen-DisteÌche, M., FreÌre, J.-M., Nicholas, R. A., and Pratt, R. F. (2003) On the substrate specificity of bacterial dd -peptidases: Evidence from two series of peptidoglycan-mimetic peptides Biochem. J. 373, 949-955
Johnson, D. A. (1953) Carboxy derivatives of benzylpenicillin J. Am. Chem. Soc. 75, 3636-3637
Xu, Y., Soto, G., Adachi, H., Van der Linden, M. P. G., Keck, W., and Pratt, R. F. (1994) Relative specificities of a series of β-lactam- reognizing enzymes towards the side chains of penicillins and acyclic thioldepsipeptides Biochem. J. 302, 851-856
Adediran, S. A., Kumar, I., Nagarajan, R., Sauvage, E., and Pratt, R. F. (2011) Kinetics of reactions of the Actinomadura R39 dd -peptidase with specific substrates Biochemistry 50, 367-387
Kuzmic, P. (1996) Program DYNAFIT for the analysis of enzyme kinetic data: Application to HIV proteinase Anal. Biochem. 237, 260-273
Duez, C., Zervosen, A., Teller, N., Melkonian, R., Banzubazé, E., Bouillenne, F., Luxen, A., and FreÌre, J.-M. (2009) Characterization of the proteins encoded by the Bacillus subtilis yoxA-dacC operon FEMS Microbiol. Lett. 300, 42-47
Shimshock, S. S., Waltermire, R. E., and DeShong, P. (1991) A total synthesis of (±)-tirandamycin B J. Am. Chem. Soc. 113, 8791-8796
Kumar, I. and Pratt, R. F. (2005) Transpeptidation of a specific substrate catalyzed by the Streptomyces R61 dd -peptidase: Characterization of a chromogenic substrate and acyl acceptor design Biochemistry 44, 9971-9979
Tipper, D. J. and Strominger, J. L. (1965) Mechanism of action of penicillins: A proposal based on their structural similarity to acyl- d -alanyl- d -alanine Proc. Natl. Acad. Sci. U.S.A. 54, 1133-1141
Pratt, R. F. (2002) Functional evolution of the serine β-lactamase active site J. Chem. Soc., Perkin Trans. 2, 851-861
Silvaggi, N. R., Josephine, H. R., Kuzin, A. P., Nagarajan, R., Pratt, R. F., and Kelly, J. A. (2005) Crystal structures of complexes between the R61 dd -peptidase and peptidoglycan-mimetic β-lactams: A non-covalent complex with a "perfect penicillin" J. Mol. Biol. 345, 521-533
Holysz, R. P. and Stavely, H. E. (1950) Carboxy derivatives of benzylpenicillin J. Am. Chem. Soc. 72, 4760-4763
Varetto, L., DeMeester, F., Monnaie, D., Marchand-Brynaert, J., Dive, G., Jacob, F., and FreÌre, J.-M. (1991) The importance of the negative charge of β-lactam compounds in the interactions with active-site serine dd -peptidases and β-lactamases Biochem. J. 278, 801-807
Curley, K. C. and Pratt, R. F. (1997) Effectiveness of tetrahedral adducts as transition-state analogs and inhibitors of the class C β-lactamase of Enterobacter cloacae P99 J. Am. Chem. Soc. 119, 1529-1538
Sauvage, E., Herman, R., Petrella, S., Duez, C., Bouillenne, F., FreÌre, J.-M., and Charlier, P. (2005) Crystal structure of the Actinomadura R39 dd -peptidase reveals new domains in penicillin binding proteins J. Biol. Chem. 280, 31249-31256
Silvaggi, N. R., Anderson, J. W., Brinsmade, S. A., Pratt, R. F., and Kelly, J. A. (2003) The crystal structure of phosphonate-inhibited d -Ala- d -Ala peptidase reveals an analogue of a tetrahedral transition state Biochemistry 42, 1199-1208
Hughes, R. C. (1970) Autolysis of isolated cell walls of Bacillus licheniformis N.C.T.C.6346 and Bacillus subtilis Marburg strain 168. Separation of the products and characterization of the mucopeptide fragments Biochem. J. 119, 849-860
Hirotsu, K., Goto, M., Okamoto, A., and Miyahara, I. (2005) Dual substrate recognition of aminotransferases Chem. Rec. 5, 160-172
Bullock, T. L., Uter, N., Nissan, T. A., and Perona, J. J. (2003) Amino acid discrimination by a class I aminoacyl-tRNA synthetase specified by negative determinants J. Mol. Biol. 328, 395-408
Chen, Y., Zhang, W., Shi, Q., Hesek, D., Lee, M., Mobashery, S., and Shoichet, B. K. (2009) Crystal structures of penicillin-binding protein 6 from Escherichia coli J. Am. Chem. Soc. 131, 14345-15354
Josephine, H. R., Charlier, P., Davies, C., Nicholas, R. A., and Pratt, R. F. (2006) Reactivity of penicillin-binding proteins with peptidoglycan- mimetic β-lactams: What's wrong with these enzymes? Biochemistry 45, 15873-15883
