title: Crystal structure and mutational analysis of aminoacylhistidine dipeptidase from Vibrio alginolyticus reveal a new architecture of M20 metallopeptidases.
abstract: Aminoacylhistidine dipeptidases (PepD, EC 3.4.13.3) belong to the family of M20 metallopeptidases from the metallopeptidase H clan that catalyze a broad range of dipeptide and tripeptide substrates, including L-carnosine and L-homocarnosine. Homocarnosine has been suggested as a precursor for the neurotransmitter γ-aminobutyric acid (GABA) and may mediate the antiseizure effects of GABAergic therapies. Here, we report the crystal structure of PepD from Vibrio alginolyticus and the results of mutational analysis of substrate-binding residues in the C-terminal as well as substrate specificity of the PepD catalytic domain-alone truncated protein PepD(CAT). The structure of PepD was found to exist as a homodimer, in which each monomer comprises a catalytic domain containing two zinc ions at the active site center for its hydrolytic function and a lid domain utilizing hydrogen bonds between helices to form the dimer interface. Although the PepD is structurally similar to PepV, which exists as a monomer, putative substrate-binding residues reside in different topological regions of the polypeptide chain. In addition, the lid domain of the PepD contains an "extra" domain not observed in related M20 family metallopeptidases with a dimeric structure. Mutational assays confirmed both the putative di-zinc allocations and the architecture of substrate recognition. In addition, the catalytic domain-alone truncated PepD(CAT) exhibited substrate specificity to l-homocarnosine compared with that of the wild-type PepD, indicating a potential value in applications of PepD(CAT) for GABAergic therapies or neuroprotection.
authors.affiliation: Department of Biological Science and Technology, National Chiao Tung University, Hsinchu 30010, Taiwan.
authors.forename: Chin-Yuan
authors.initials: CY
authors.lastname: Chang
authors.forename: Yin-Cheng
authors.initials: YC
authors.lastname: Hsieh
authors.forename: Ting-Yi
authors.initials: TY
authors.lastname: Wang
authors.forename: Yi-Chin
authors.initials: YC
authors.lastname: Chen
authors.forename: Yu-Kuo
authors.initials: YK
authors.lastname: Wang
authors.forename: Ting-Wei
authors.initials: TW
authors.lastname: Chiang
authors.forename: Yi-Ju
authors.initials: YJ
authors.lastname: Chen
authors.forename: Cheng-Hsiang
authors.initials: CH
authors.lastname: Chang
authors.forename: Chun-Jung
authors.initials: CJ
authors.lastname: Chen
authors.forename: Tung-Kung
authors.initials: TK
authors.lastname: Wu
mesh_terms: D000596:Amino Acids / Q000737:chemistry; D020134:Catalytic Domain; D018360:Crystallography, X-Ray / Q000379:methods; D004252:DNA Mutational Analysis / Q000379:methods; D004150:Dipeptidases / Q000737:chemistry*; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D006860:Hydrogen Bonding; D007700:Kinetics; D008968:Molecular Conformation; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D044149:Vibrio alginolyticus / Q000201:enzymology*
chemical_list: D000596:Amino Acids; D004150:Dipeptidases; C019673:aminoacyl-histidine dipeptidase
references.citation: Acta Crystallogr D Biol Crystallogr. 2006 Jan;62(Pt 1):72-82
references.pmid: 16369096
references.citation: J Biol Chem. 2003 Nov 7;278(45):44496-504
references.pmid: 12933810
references.citation: Biochemistry (Mosc). 2000 Jul;65(7):766-70
references.pmid: 10951093
references.citation: Methods Enzymol. 1997;276:307-26
references.pmid: 27754618
references.citation: J Biol Chem. 2008 Oct 3;283(40):27289-99
references.pmid: 18550540
references.citation: Diabetes. 2007 Oct;56(10):2425-32
references.pmid: 17601992
references.citation: Structure. 1997 Mar 15;5(3):337-47
references.pmid: 9083113
references.citation: Acta Crystallogr D Biol Crystallogr. 2004 Dec;60(Pt 12 Pt 1):2126-32
references.pmid: 15572765
references.citation: J Biol Chem. 2003 Dec 19;278(51):51851-62
references.pmid: 14534321
references.citation: J Neurochem. 1982 Jan;38(1):242-5
references.pmid: 7108529
references.citation: Structure. 2002 Aug;10(8):1097-106
references.pmid: 12176387
references.citation: J Mol Biol. 2007 Apr 27;368(2):450-63
references.pmid: 17362992
references.citation: Neurology. 1972 Jun;22(6):644-55
references.pmid: 4673339
references.citation: J Int Neuropsychol Soc. 2006 May;12(3):416-23
references.pmid: 16903134
references.citation: Neurology. 1999 Feb;52(3):473-8
references.pmid: 10025774
references.citation: Curr Med Chem. 2005;12(20):2293-315
references.pmid: 16181134
references.citation: Proteins. 2003 Feb 15;50(3):437-50
references.pmid: 12557186
references.citation: Structure. 1994 Apr 15;2(4):283-91
references.pmid: 8087555
references.citation: Eur J Biochem. 1985 May 2;148(3):447-53
references.pmid: 3838935
references.citation: FEBS J. 2008 Oct;275(20):5007-20
references.pmid: 18783432
references.citation: J Mol Biol. 1997 Feb 7;265(5):620-36
references.pmid: 9048953
references.citation: Neurology. 2001 Mar 27;56(6):709-15
references.pmid: 11274303
references.citation: Leuk Lymphoma. 2002 Nov;43(11):2139-43
references.pmid: 12533039
references.citation: Gene. 1984 Nov;31(1-3):31-8
references.pmid: 6396165
references.citation: Microbiology (Reading). 1994 Oct;140 ( Pt 10):2591-600
references.pmid: 7528082
references.citation: Acta Crystallogr Sect F Struct Biol Cryst Commun. 2009 Mar 1;65(Pt 3):216-8
references.pmid: 19255468
references.citation: J Biol Chem. 2003 Feb 21;278(8):6521-31
references.pmid: 12473676
references.citation: Biochemistry (Mosc). 2000 Jul;65(7):751-6
references.pmid: 10951091
references.citation: J Biochem Mol Toxicol. 2000;14(4):215-20
references.pmid: 10789500
references.citation: J Bacteriol. 1997 Jun;179(11):3410-5
references.pmid: 9171382
references.citation: J Neurosci Res. 2002 May 15;68(4):463-9
references.pmid: 11992473
references.citation: J Chromatogr A. 2004 Mar 26;1031(1-2):67-78
references.pmid: 15058568
references.citation: Int J Biochem Cell Biol. 1998 Aug;30(8):863-8
references.pmid: 9744078
references.citation: FEMS Microbiol Lett. 1994 Oct 15;123(1-2):153-9
references.pmid: 7988883
references.citation: Comp Biochem Physiol B Biochem Mol Biol. 2001 Mar;128(3):469-81
references.pmid: 11250542
references.citation: Pediatr Res. 1973 Jul;7(7):601-6
references.pmid: 4718759
references.citation: N Engl J Med. 1967 Dec 7;277(23):1219-27
references.pmid: 6058610
references.citation: J Med Chem. 1999 Mar 25;42(6):951-6
references.pmid: 10090777
references.citation: Infect Immun. 1997 Oct;65(10):4158-64
references.pmid: 9317022
references.citation: Lancet. 1968 Jun 8;1(7554):1229-30
references.pmid: 4172777
references.citation: J Phys Chem B. 2008 Feb 28;112(8):2494-500
references.pmid: 18247603
references.citation: Eur J Biochem. 2002 Jan;269(2):443-50
references.pmid: 11856302