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Article: Mechanism of 8-amino-7-oxononanoate synthase: Spectroscopic, kinetic, and crystallographic studies

TitleMechanism of 8-amino-7-oxononanoate synthase: Spectroscopic, kinetic, and crystallographic studies
Authors
Issue Date2000
PublisherAmerican Chemical Society. The Journal's web site is located at http://pubs.acs.org/biochemistry
Citation
Biochemistry, 2000, v. 39 n. 3, p. 516-528 How to Cite?
Abstract8-Amino-7-oxononanoate synthase (also known as 7-keto-8-aminopelargonate synthase, EC 2.3.1.47) is a pyridoxal 5'-phosphate-dependent enzyme which catalyzes the decarboxylative condensation of L-alanine with pimeloyl-CoA in a stereospecific manner to form 8(S)-amino-7-oxononanoate. This is the first committed step in biotin biosynthesis. The mechanism of Escherichia coli AONS has been investigated by spectroscopic, kinetic, and crystallographic techniques. The X-ray structure of the holoenzyme has been refined at a resolution of 1.7 Å (R = 18.6%, R(free) = 21.2%) and shows that the plane of the imine bond of the internal aldimine deviates from the pyridine plane. The structure of the enzyme-product external aldimine complex has been refined at a resolution of 2.0 Å (R = 21.2%, R(free) = 27.8%) and shows a rotation of the pyridine ring with respect to that in the internal aldimine, together with a significant conformational change of the C-terminal domain and subtle rearrangement of the active site hydrogen bonding. The first step in the reaction, L-alanine external aldimine formation, is rapid (k1 = 2 x 104 M- 1 s-1). Formation of an external aldimine with D-alanine, which is not a substrate, is significantly slower (k1 = 125 M-1 s-1). Binding of D- alanine to AONS is enhanced approximately 2-fold in the presence of pimeloyl- CoA. Significant substrate quinonoid formation only occurs upon addition of pimeloyl-CoA to the preformed L-alanine external aldimine complex and is preceded by a distinct lag phase (~30 ms) which suggests that binding of the pimeloyl-CoA causes a conformational transition of the enzyme external aldimine complex. This transition, which is inferred by modeling to require a rotation around the Cα-N bond of the external aldimine complex, promotes abstraction of the Cα proton by Lys236. These results have been combined to form a detailed mechanistic pathway for AONS catalysis which may be applied to the other members of the α-oxoamine synthase subfamily.
Persistent Identifierhttp://hdl.handle.net/10722/154134
ISSN
2015 Impact Factor: 2.876
2015 SCImago Journal Rankings: 1.769
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorWebster, SPen_US
dc.contributor.authorAlexeev, Den_US
dc.contributor.authorCampopiano, DJen_US
dc.contributor.authorWatt, RMen_US
dc.contributor.authorAlexeeva, Men_US
dc.contributor.authorSawyer, Len_US
dc.contributor.authorBaxter, RLen_US
dc.date.accessioned2012-08-08T08:23:26Z-
dc.date.available2012-08-08T08:23:26Z-
dc.date.issued2000en_US
dc.identifier.citationBiochemistry, 2000, v. 39 n. 3, p. 516-528en_US
dc.identifier.issn0006-2960en_US
dc.identifier.urihttp://hdl.handle.net/10722/154134-
dc.description.abstract8-Amino-7-oxononanoate synthase (also known as 7-keto-8-aminopelargonate synthase, EC 2.3.1.47) is a pyridoxal 5'-phosphate-dependent enzyme which catalyzes the decarboxylative condensation of L-alanine with pimeloyl-CoA in a stereospecific manner to form 8(S)-amino-7-oxononanoate. This is the first committed step in biotin biosynthesis. The mechanism of Escherichia coli AONS has been investigated by spectroscopic, kinetic, and crystallographic techniques. The X-ray structure of the holoenzyme has been refined at a resolution of 1.7 Å (R = 18.6%, R(free) = 21.2%) and shows that the plane of the imine bond of the internal aldimine deviates from the pyridine plane. The structure of the enzyme-product external aldimine complex has been refined at a resolution of 2.0 Å (R = 21.2%, R(free) = 27.8%) and shows a rotation of the pyridine ring with respect to that in the internal aldimine, together with a significant conformational change of the C-terminal domain and subtle rearrangement of the active site hydrogen bonding. The first step in the reaction, L-alanine external aldimine formation, is rapid (k1 = 2 x 104 M- 1 s-1). Formation of an external aldimine with D-alanine, which is not a substrate, is significantly slower (k1 = 125 M-1 s-1). Binding of D- alanine to AONS is enhanced approximately 2-fold in the presence of pimeloyl- CoA. Significant substrate quinonoid formation only occurs upon addition of pimeloyl-CoA to the preformed L-alanine external aldimine complex and is preceded by a distinct lag phase (~30 ms) which suggests that binding of the pimeloyl-CoA causes a conformational transition of the enzyme external aldimine complex. This transition, which is inferred by modeling to require a rotation around the Cα-N bond of the external aldimine complex, promotes abstraction of the Cα proton by Lys236. These results have been combined to form a detailed mechanistic pathway for AONS catalysis which may be applied to the other members of the α-oxoamine synthase subfamily.en_US
dc.languageengen_US
dc.publisherAmerican Chemical Society. The Journal's web site is located at http://pubs.acs.org/biochemistryen_US
dc.relation.ispartofBiochemistryen_US
dc.subject.meshAcyl Coenzyme A - Metabolismen_US
dc.subject.meshAcyltransferases - Chemistry - Metabolismen_US
dc.subject.meshAlanine - Metabolismen_US
dc.subject.meshAmino Acid Sequenceen_US
dc.subject.meshBacillus - Enzymologyen_US
dc.subject.meshBinding Sitesen_US
dc.subject.meshCrystallography, X-Rayen_US
dc.subject.meshEscherichia Coli - Enzymologyen_US
dc.subject.meshHydrogen Bondingen_US
dc.subject.meshKineticsen_US
dc.subject.meshMass Spectrometryen_US
dc.subject.meshModels, Molecularen_US
dc.subject.meshMolecular Sequence Dataen_US
dc.subject.meshProtein Conformationen_US
dc.subject.meshSpectrophotometryen_US
dc.subject.meshSubstrate Specificityen_US
dc.titleMechanism of 8-amino-7-oxononanoate synthase: Spectroscopic, kinetic, and crystallographic studiesen_US
dc.typeArticleen_US
dc.identifier.emailWatt, RM:rmwatt@hku.hken_US
dc.identifier.authorityWatt, RM=rp00043en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.doi10.1021/bi991620jen_US
dc.identifier.pmid10642176-
dc.identifier.scopuseid_2-s2.0-0034711757en_US
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-0034711757&selection=ref&src=s&origin=recordpageen_US
dc.identifier.volume39en_US
dc.identifier.issue3en_US
dc.identifier.spage516en_US
dc.identifier.epage528en_US
dc.identifier.isiWOS:000084990800004-
dc.publisher.placeUnited Statesen_US
dc.identifier.scopusauthoridWebster, SP=7101874218en_US
dc.identifier.scopusauthoridAlexeev, D=6701529096en_US
dc.identifier.scopusauthoridCampopiano, DJ=6602680734en_US
dc.identifier.scopusauthoridWatt, RM=7102907536en_US
dc.identifier.scopusauthoridAlexeeva, M=35607988700en_US
dc.identifier.scopusauthoridSawyer, L=7102690704en_US
dc.identifier.scopusauthoridBaxter, RL=26643014900en_US

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