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Structure-function studies of beta-lactam biosynthetic enzymes

Öster, Linda (2005). Structure-function studies of beta-lactam biosynthetic enzymes. Diss. (sammanfattning/summary) Uppsala : Sveriges lantbruksuniv., Acta Universitatis Agriculturae Sueciae, 1652-6880 ; 2005:1
ISBN 91-576-7000-5
[Doctoral thesis]

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Beta-lactam compounds belong to the most important antibiotics in current use. The increasing occurrence of bacterial resistance against antibiotics, which threatens to move us back to pre-antibiotic era, calls for the development of new antibiotics. Beta-lactams are produced by fermentation in the microorganism, therefore knowledge about the biosynthetic enzymes involved is vital for the production of new antibiotics to escape resistance. This thesis presents structure-function studies on enzymes involved in the biosynthesis of the beta-lactam antibiotics cephalosporins and cephamycins DAOCS is a non-heme Fe(II) dioxygenase and catalyses the oxidative ring expansion of the penicillin nucleus into the nucleus of cephalosporins. Structures of DAOCS with substrates, product and cofactors obtained from two different crystal forms are presented here. The structural results suggest a mechanism for cephalosporin formation where 2-oxoglutarate and dioxygen need to react first to produce an oxidizing iron species, followed by reaction with the penicillin substrate. Structural differences in the two crystal forms indicate conformational flexibility in the C terminus and point to a role for the C terminus in catalysis. Biosynthesis of cephamycins (C7-methoxylated cephalosporins) is catalysed by two enzymes, cmcI and cmcJ, but the details of catalysis are largely unknown. The crystal structure of cmcI, presented here, is a hexamer consisting of a C-terminal Rossman domain and a smaller N-terminal domain. The N-terminal domain is involved in oligomerisation and the Rossmann domain binds SAM and SAH in a fashion common for methyltransferases, with a bound magnesium ion in the vicinity of SAM. The expected cephalosporin binding site is occupied by PEG, which ligates the magnesium ion. From docking studies of a cephalosoporin molecule to the cmcI-Mg2+-SAM structure, a model for substrate binding is proposed. Altogether, the results suggest cmcI is a methyltransferase that catalyses the second catalytic step in cephamycin biosynthesis.

Authors/Creators:Öster, Linda
Title:Structure-function studies of beta-lactam biosynthetic enzymes
Series Name/Journal:Acta Universitatis Agriculturae Sueciae
Year of publishing :January 2005
Number of Pages:73
ALLI. Valegård, K., Terwisscha van Scheltinga, A.C., Dubus, A., Ranghino, G., Öster, L.M., Hajdu, J. & Andersson, I. (2004) The structural basis of cephalosporin formation in a mononuclear ferrous enzyme. Nat. Struct. Mol. Biol. 11, 95-101. II. Öster, L.M., Terwisscha van Scheltinga, A.C., Valegård, K., MacKenzie Hose, A., Dubus, A., Hajdu, J. & Andersson, I. (2004) Conformational flexibility of the C Terminus with Implications for Substrate Binding and Catalysis Revealed in a New Crystal Form of Deacetoxycephalosporin C Synthase. J. Mol. Biol. 343, 157-171. III. Lester, D.R., Öster, L.M., Svenda, M. & Andersson, I. (2004) Expression, purification, crystallization and preliminary X-ray diffraction studies of the cmcI component of Streptomyces clavuligerus 7a-cephem-methoxylase. Acta Crystallogr. D 60, 1618-1621. IV. Öster, L.M., Lester, D.R., Terwisscha van Scheltinga, A.C., Svenda M., Généreux, C. & Andersson, I. (2004) Insights into Cephamycin Biosynthesis: the Crystal Structure of cmcI from Streptomyces clavuligerus. In Manuscript
Place of Publication:Uppsala
ISBN for printed version:91-576-7000-5
Publication Type:Doctoral thesis
Full Text Status:Public
Agris subject categories.:X Agricola extesions > X30 Life sciences
Subjects:Not in use, please see Agris categories
Agrovoc terms:molecular biology
Keywords:beta-lactam antibiotics, 2-oxoglutarate dependent dioxygenases, cephamycin biosynthesis, protein crystallography, Streptomyces clavuligerus, deacetoxycephalosporin C Synthase, cmcI
Permanent URL:
ID Code:737
Department:(NL, NJ) > Dept. of Molecular Biology (until 131231)
Deposited By: Linda Öster
Deposited On:26 Jan 2005 00:00
Metadata Last Modified:02 Dec 2014 10:06

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