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The Enolization Chemistry of a Thioester-Dependent Racemase: The 1.4 angstrom Crystal Structure of a Reaction Intermediate Complex Characterized by Detailed QM/MM Calculations

  1. Author:
    Sharma, S.
    Bhaumik, P.
    Schmitz, W.
    Venkatesan, R.
    Hiltunen, J. K.
    Conzelmann, E.
    Juffer, A. H.
    Wierenga, R. K.
  2. Author Address

    [Sharma, Satyan; Venkatesan, Rajaram; Hiltunen, J. Kalervo; Juffer, Andre H.; Wierenga, Rik K.] Univ Oulu, Bioctr Oulu, FI-90014 Oulu, Finland. [Sharma, Satyan; Venkatesan, Rajaram; Hiltunen, J. Kalervo; Juffer, Andre H.; Wierenga, Rik K.] Univ Oulu, Dept Biochem, FI-90014 Oulu, Finland. [Bhaumik, Prasenjit] NCI, Prot Struct Sect, Macromol Crystallog Lab, Frederick, MD 21702 USA. [Schmitz, Werner; Conzelmann, Ernst] Univ Wurzburg, Theodor Boveri Inst Biowissensch, Biozentrum, D-97074 Wurzburg, Germany.;Wierenga, RK (reprint author), Univ Oulu, Bioctr Oulu, POB 3000, FI-90014 Oulu, Finland;rik.wierenga@oulu.fi
    1. Year: 2012
    2. Date: Mar
  1. Journal: Journal of Physical Chemistry B
    1. 116
    2. 11
    3. Pages: 3619-3629
  2. Type of Article: Article
  3. ISSN: 1520-6106
  1. Abstract:

    In the active site of the bacterial alpha-methylacyl-CoA racemase of Mycobacterium tuberculosis (MCR), the chirality of the 2-methyl branched C2-atom is interconverted between (S) and (R) isomers. Protein crystallographic data and quantum mechanics/molecular mechanics (QM/MM) computational approaches show that this interconversion is achieved via a planar enolate intermediate. The crystal structure, at 1.4 angstrom, visualizes the mode of binding of a reaction intermediate analogue, 2-methylacetoacetyl-CoA, in a well-defined planar enolate form. The computational studies confirm that in the conversion from (S) to (R), first a proton is abstracted by N delta 1 (His126), and subsequently the planar enolate form is reprotonated by O delta 2 (Asp156). The calculations also show that the negatively charged thioester oxygen of the enolate intermediate is stabilized by an oxyanion hole formed by N (Asp127), as well as by the side chain atoms of the catalytic residues, Asp 156 and His 126, both being protonated simultaneously, at the intermediate stage of the catalytic cycle. The computational analysis also reveals that the conversion of the (S)- to (R)- chirality is achieved by a movement of 1.7 angstrom of the chiral C2-carbon, with smaller shifts (approximately 1 angstrom) of the carbon atom of the 2-methyl group, the C3-atom of the fatty acid tail, and the C1-carbon and O1-oxygen atoms of the thioester moiety.

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External Sources

  1. DOI: 10.1021/jp210185m
  2. WOS: 000301766700015

Library Notes

  1. Fiscal Year: FY2011-2012
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