The trans effect of residue 408 in protocatechuate 3,4-dioxygenase

M. P. Valley, N. Klang, D. H. Ohlendorf, J. I. Lipscornb

Research output: Contribution to journalArticlepeer-review

Abstract

Protncatechuate. :t, 1-dioxygenase catafy/es the intradiol ring cleavage of 3.1 dihydroxybenzuic acid with The incorporation of both atoms of 0?. Spectio s(0pic and < rystallographic data have established that a single l-Vi+ is bound in the active site by two lyr, two His. and a single OH- . In the proposed mechani.-rri. the substrate initially chelates the Ke3+ and maintains the 0-cooniinate geometry by displacing the OH" and axial Tyr ligands. Two of the remaining l''e !4" ligands exert Irans effects on the nubs t rate, leading to asymmetry in bind ing of the hydroxvU; this activates the substrat"1 by inducing the weaker binding i Oil to ketonize and accumulate charge at carbon- 1 \vhere 0-2 attacks. The ligand opposite the II-O1I. the equatorial l'yr -108. has been replaced, through sih'-diiected inntagenesis, with a (ihi to test its role in th'1 catalytic mechanism. Model studies suggest tiiat an increase in Lewis acidity engendered by a I'vr to (ilu mutation at the metal center should yield an increase in (atalyti< artiviu. However, the weaker Irans effect of a carboxylic acid, compared to .1 pheiiolate. should strengthen the bond between the Fe1+ and :J-()H so as to hinder ketonization and thus inhibit catalysis. Y408F is observed to bind IV' .ind substrate luit has no detectable activity, suggesting that trans effects rna\ ptd\ i more important role than Lewis acidity in the catalytic mechanism, binding assays and structural analysis are in progress. '.Work supported by Mil tirant (IM24689.).

Original languageEnglish (US)
Pages (from-to)A1309
JournalFASEB Journal
Volume11
Issue number9
StatePublished - Dec 1 1997

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