Sterol C24-methyltransferase: Physio- and stereo-chemical features of the sterol C3 group required for catalytic competence

Alicia L. Howard, Jialin Liu, Gamal A. Elmegeed, Emily K. Collins, Kalgi S. Ganatra, Chizaram A. Nwogwugwu, W. David Nes

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

Sterol C24-methyltransferases (24-SMTs) catalyze the electrophilic alkylation of Δ24-sterols to a variety of sterol side chain constructions, and the C3- moiety is the primary determinant for substrate binding by these enzymes. To determine what specific structural features of the C3-polar group ensure sterol catalysis, a series of structurally related C3-analogs of lanosterol that differed in stereochemistry, bulk and electronic properties were examined against the fungal 24-SMT from Paracoccidioides brasiliensis (Pb) which recognize lanosterol as the natural substrate. Analysis of the magnitude of sterol C24-methylation activity (based on the kinetic constants of Vmax/Km and product distributions determined by GC-MS) resulting from changes at the C3-position in which the 3β-OH was replaced by 3α-OH, 3β-acetyl, 3-oxo, 3-OMe, 3β-F, 3β-NH2 (protonated species) or 3H group revealed that lanosterol and five substrate analogs were catalyzed and yielded identical side chain products whereas neither the 3H- or 3α-OH lanosterol derivatives were productively bound. Taken together, our results demonstrate a chemical complementarity involving hydrogen bonding formation of specific active site contacts to the nucleophilic C3-group of sterol is required for proper orientation of the substrate C-methyl intermediate in the activated complex.

Original languageEnglish
Pages (from-to)43-50
Number of pages8
JournalArchives of Biochemistry and Biophysics
Volume521
Issue number1-2
DOIs
StatePublished - May 2012

Keywords

  • Fluorinated sterol
  • Hydrogen bonding
  • Lanosterol
  • Paracoccidioides brasiliensis
  • Sterol C24-methyltransferase
  • Sterol catalysis

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