Stability, denaturation and refolding of Mycobacterium tuberculosis MfpA, a DNA mimicking protein that confers antibiotic resistance

Sergei Khrapunov, Michael Brenowitz

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

MfpA from Mycobacterium tuberculosis is a founding member of the pentapeptide repeat class of proteins (PRP) that is believed to confer bacterial resistance to the drug fluoroquinolone by mimicking the size, shape and surface charge of duplex DNA. We show that phenylalanine side chain stacking stabilizes the N-terminus of MfpA's pentapeptide thus extending the DNA mimicry analogy. The Lumry-Eyring model was applied to multiple spectral measures of MfpA denaturation revealing that the MfpA dimer dissociates to monomers which undergo a structural transition that leads to aggregation. MfpA retains high secondary and tertiary structure content under denaturing conditions. Dimerization stabilizes MfpA's pentapeptide repeat fold. The high Arrhenius activation energy of the barrier to aggregate formation rationalizes its stability. The mechanism of MfpA denaturation and refolding is a 'double funnel' energy landscape where the 'native' and 'aggregate' funnels are separated by the high barrier that is not overcome during in vitro refolding.

Original languageEnglish (US)
Pages (from-to)33-40
Number of pages8
JournalBiophysical Chemistry
Volume159
Issue number1
DOIs
StatePublished - Nov 2011

Keywords

  • Antibiotic resistance
  • CD
  • Fluorescence
  • MfpA
  • Pentapeptide repeat protein
  • Protein folding

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Organic Chemistry

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