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    Table of contents
    1. 1. Protein Summary
    2. 2. Ligand Summary

    Title Crystal structure of putative pyridoxal 5'-phosphate-dependent C-S lyase (YP_813084.1) from LACTOBACILLUS DELBRUECKII BULGARICUS ATCC BAA-365 at 1.61 A resolution. To be published
    Site JCSG
    PDB Id 3dzz Target Id 377395
    Molecular Characteristics
    Source Lactobacillus delbrueckii subsp. bulgaricus atcc baa-365
    Alias Ids TPS17306,YP_813084.1, 3.40.640.10, 335798 Molecular Weight 43992.97 Da.
    Residues 390 Isoelectric Point 5.02
    Sequence maekqydfthvpkrqgnsikwgvlkekelpmwiaemdfkiapeimasmeeklkvaafgyesvpaeyyka vadweeiehrarpkedwcvfasgvvpaisamvrqftspgdqilvqepvynmfysviegngrrvissdli yenskysvnwadleeklatpsvrmmvfcnphnpigyawseeevkriaelcakhqvllisdeihgdlvlt deditpaftvdwdaknwvvslispsktfnlaalhaacaiipnpdlraraeesfflagigepnllaipaa iaayeeghdwlrelkqvlrdnfayareflakevpevkvldsnasylawvdisalgmnaedfckylrekt gliisagngyrgnghefvrinlacpkelvidgmqrlkqgvlnlnn
      BLAST   FFAS

    Structure Determination
    Method XRAY Chains 2
    Resolution (Å) 1.61 Rfree 0.175
    Matthews' coefficent 2.06 Rfactor 0.144
    Waters 575 Solvent Content 40.40

    Ligand Information



    Protein Summary

    The LBUL1103 (patC) gene from Lactobacillus delbrueckii bulgaricus atcc baa-365 encodes a PLP-dependent aminotransferase (PF00155, COG1168, EC 2.6.1.-). The structure contains two domains and adopts a PLP-dependent transferase-like fold with significant structural similarity to cystalysins (PDB ids 1C7N, 1D2F), aspartate aminotransferases (PDB ids 1J32, 1O4S) and other aminotransferases of varying specificities. A search with PSI-BLAST indicates that the protein is likely as a beta-cystathionase. Previous work has shown that this enzyme can have a dual role, functioning both as a beta-cystathionase and as a modulator of maltose gene expression [Ref].


    To do: identify PLP site, catalytic residues, ligand?


    Kinetic assays establish that 3DZZ is a cystathionine β-lyase


    The gene for protein YP_813084.1 (patC) from Lactobacillus delbruecki subsp. bulgaricus was identified to encode a carbon-sulfur lyase (C-S lyase), specifically cystathionine β-lysase (LdCBL; EC, in part because of its ability to complement methionine auxotrophy in an Escherichia coli metC mutant1. The crystal structure for LdCBL (PDB id 3DZZ) was determined with a resolution of 1.61 Å by the JCSG2.


    Sequence analysis indicates LdCBL contains a conserved aspartate aminotransferase (AAT) fold characteristic of the PLP-dependent AAT-superfamily3. Additionally, structural searches through VAST classify LdCBL as fold-type I4; this fold type is a well-characterized fold found in other C-S lyase proteins with similar proposed function:  E. coli cystathionine β-lysase (EcCBL); E. coli bifunctional maltose regulon repressor and cystathionine β-lysase (EcMCBL); Streptococcus anginosus βC-S lyase (SaCSL;; and cystalysin, a βC-S lyase fromTreponema denticola (TdCSL;  Each of these proteins contains a conserved catalytic lysine (Lys233 in LdCBL), which binds to PLP through a Schiff base linkage5,6,7,8..


    Sequence alignment of LdCBL with EcCBL, EcMCBL, SaCSL, and TdCSL identified conserved residues important to catalysis: Tyr59, Val93, Asn170, Ser230, Ser232 and Lys233 are important in PLP-binding and are conserved across the βC-S lysases9; Tyr118 and Asp198 are conserved residues common in transsulfuration mechanisms6 (Figure 1).


    Kinetics data substantiate the previously putative annotation of 3DZZ as a CBL in the Lactobacillus delbruecki bulgaricus transulfuration pathway. L-cystathionine (L-Cth), L-cysteine (L-Cys) and L-cysteine (L-Cys2)were tested as potential substrates for LdCBL


    LdCBL catalyzes the conversion of L-Cth (KM = 9 ±2 mM, kcat = 1.2 ±0.1 s-1, kcat/KM = 0.13 ± 0.03 s-1 mM-1) to L-homocysteine and pyruvate (Figure 2A). Substrate inhibition was observed with L-Cth above  20 µM (Figure 2A). While enzymatic activity was seen with L-Cys and L-Cys2 as substrates, accurate kinetic parameters were not able to be determined. Substrate inhibition was observed with L-Cys at higher concentrations (Figure 2B), preventing any reliable kinetic parameter determination. The relative insolubility of L-Cys2 prevented rate measurement at or near the Vmax.



    1.    Aubel D., Germond J.E., Gilbert C., and Atlan D. 2002.  Isolation of the patC gene encoding the cystathionine β-lyase of Lactobacillus delbrueckii subsp. bulgaricus and molecular analysis of inter-strain variability in enzyme biosynthesis.  Microbiology. 148: 2029-36.

    2.    Elsiger M., Deacon A.M., Godzik A., Lesley S.A., Wooley J., Wüthrich K., Wilson I.A. 2010. The JCSG high-throughput structural biology pipeline. Acta Crystallographic Sect F Structural Biology Crystal Community. 66(10): 1137-42.

    3.    Altschul S.F., Gish W., Miller W., Myers E.W. and Lipman D.J. 1990. Basic local alignment search tool. Journal of Moecular Biology.215: 403-10.

    4.    Gibrat J.F., Madej T. and Bryant S.H. 1996. Surprising similarities in structure comparison. Curr. Opin. Struct. Biol. 1996 6: 377-85.Holm L and Rosenstrӧm P. 2010. Dali server: conservation mapping in 3D. Nucleic Acids Research. 38: W545-49

    5.    Clausen T., Huber R., Laber B., Pohlenz H., and Messerschmidt A. 1996. Crystal structure of the pyridoxal-5’-phosphate dependent cystathionine β-lyase from Escherichia coli at 1.83 Å. Journal of Molecular Biology.262: 202-24.

    6.    Clausen T., Schlegel E., Peist R., Schneider E., Steegborn C., Chang Y., Haase A., Bourenkov G.P., Bartunik H.D., and Boos W. 2000. X-ray structure of MalY from Escherichia coli: a pyridoxal-5’-phosphate dependent enzyme acting as a modulator in mal gene expression. The EMBO Journal. 19: 831-42.

    7.    Kezuka Y, Yoshida Y, and Nonaka T. 2012. Structural insights into catalysis by βC-S lyase from Streptococcus anginosus. Proteins: Structure, Function, and Bioinformatics. 80: 2447-57.

    8.    Krupka H.I., Huber R., Holt S.C., and Clausen T. 2000. Crystal structure of cystalysin from Treponema denticola: a pyridoxal-5’-phosphate dependent protein acting as a haemolytic enzyme. The EMBO Journal.19: 3168-78.

    9.    Sievers F., Wilm A., Dineen D.G., Gibson T.J., Karplus K., Li W., Lopez R., McWilliam H., Remmert M., Söding J., Thompson J.D., and Higgins D.G. 2011. Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega. Molecular Systems Biology.7: 539-45.


    BioLEd Contributors: Annelise Bederman, Erik Eklund, China Green, Victoria Hall, Jordan Kramer, Katherine Lambertson, Steven Tan, Catrina Campbell, Ellen Schleckman, Cameron Mura, Carol Price, Linda Columbus. Funded by NSF DUE 1044858.

    Ligand Summary




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