title: Mass spectrometric analysis of dimer-disrupting mutations in Plasmodium triosephosphate isomerase.
abstract: Electrospray ionization mass spectrometry (ESI MS) under nanospray conditions has been used to examine the effects of mutation at two key dimer interface residues, Gln (Q) 64 and Thr (T) 75, in Plasmodium falciparum triosephosphate isomerase. Both residues participate in an intricate network of intra- and intersubunit hydrogen bonds. The gas phase distributions of dimeric and monomeric protein species have been examined for the wild type enzyme (TWT) and three mutants, Q64N, Q64E, and T75S, under a wide range of collision energies (40-160 eV). The results established the order of dimer stability as TWT > T75S > Q64E ∼ Q64N. The mutational effects on dimer stability are in good agreement with the previously reported estimates, based on the concentration dependence of enzyme activity. Additional experiments in solution, using inhibition of activity by a synthetic dimer interface peptide, further support the broad agreement between gas phase and solution studies.
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Debarati
authors.initials: D
authors.lastname: Bandyopadhyay
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Sunita
authors.initials: S
authors.lastname: Prakash
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Kallol
authors.initials: K
authors.lastname: Gupta
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India. Electronic address: pb@mbu.iisc.ernet.in.
authors.forename: Padmanabhan
authors.initials: P
authors.lastname: Balaram
mesh_terms: D000818:Animals; D019281:Dimerization; D006860:Hydrogen Bonding; D013058:Mass Spectrometry / Q000379:methods*; D009154:Mutation; D010961:Plasmodium / Q000201:enzymology*; D014305:Triose-Phosphate Isomerase / Q000235:genetics*
chemical_list: D014305:Triose-Phosphate Isomerase
keywords: Gas phase dimers; Protein dimer stability; Protein mass spectrometry; Triosephosphate isomerase; Triosephosphate isomerase interface mutants