Detection of the protein dimers, multiple monomeric states and hydrated forms of Plasmodium falciparum triosephosphate isomerase in the gas phase.
Dimeric and monomeric forms of the enzyme triosephosphate isomerase (TIM) from Plasmodium falciparum (Pf) have been detected under conditions of nanoflow by electrospray mass spectrometry. The dimer (M = 55 663 Da) exhibits a narrow charge state distribution with intense peaks limited to values of 18(+) to 21(+), maximal intensity being observed for charge states 19(+) and 20(+). A monomeric species with a charge state distribution ranging from 11(+) to 16(+) is also observed, which may be assigned to folded dissociated subunits. Complete dimer dissociation results under normal electrospray condition. The effects of solution pH and source temperature have been investigated. The observation of four distinct charge state distributions which may be assigned to a dimer, folded monomer, partially folded monomer and unfolded monomer is reported. Circular dichromism and fluorescence studies of Pf TIM at low pH support the retention of substantial secondary and tertiary structures. Satellite peaks in mass spectra corresponding to hydrated species are also observed and isotope shift upon deuteration is demonstrated. The analysis of all available independent crystal structures of Pf TIM and TIMs from other organisms permits identification of structurally conserved water molecules. Hydration observed in the dimer and folded monomeric forms in the gas phase may correspond to these conserved sites.
Thakur
Suman S
SS
Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.
Deepalakshmi
P D
PD
Gayathri
P
P
Banerjee
Mousumi
M
Murthy
M R N
MR
Balaram
P
P
D000818:Animals; D002942:Circular Dichroism; D003461:Crystallography; D019281:Dimerization; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000235:genetics; D013058:Mass Spectrometry; D008958:Models, Molecular; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000235:genetics; D011487:Protein Conformation; D013696:Temperature; D014305:Triose-Phosphate Isomerase / Q000235:genetics* / Q000378:metabolism*
D007527:Isoenzymes; D014305:Triose-Phosphate Isomerase