The effects of sodium perchlorate on rabbit muscle enolase--Spectral characterization of the monomer.
Incubation of rabbit beta beta enolase in NaClO4 (< or = O.3 M) results in a loss of enzymatic activity and striking changes in the second-derivative ultraviolet spectrum of enolase. HPLC gel filtration shows that dissociation of the dimeric enzyme is occurring. We have used molecular modelling, fluorescence and circular dichroic spectroscopy to examine the structural differences between the monomeric and dimeric forms of this protein. In the dimer, the tyrosine residues are in a non-polar environment; upon dissociation, two of them that were at the dimer interface become exposed. This results in large changes in the second-derivative spectrum. Both the tryptophan fluorescence emission spectrum and the aromatic region of the CD spectrum indicate that there are also changes in the environment of other aromatic residues. No perturbations in the peptide bond region of the CD spectrum are observed. We propose that the major structural effect of NaClO4 is to increase the flexibility of the loops connecting the helices and strands of the alpha/beta barrel of enolase. These loops, which contain about half of the aromatic residues, contain some of the residues of the active site and other residues involved in subunit contacts. Increased flexibility of the loops could disrupt both subunit interactions and the structure of the active site.
Kornblatt
M J
MJ
Enzyme Research Group, Department of Chemistry and Biochemistry, Concordia University, Montreal, Canada.
Al-Ghanim
A
A
Kornblatt
J A
JA
D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D002942:Circular Dichroism; D007527:Isoenzymes / Q000187:drug effects*; D009132:Muscles / Q000201:enzymology*; D010472:Perchlorates / Q000494:pharmacology*; D010751:Phosphopyruvate Hydratase / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D011487:Protein Conformation; D011817:Rabbits; D017670:Sodium Compounds / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet
D000596:Amino Acids; D007527:Isoenzymes; D010472:Perchlorates; D017670:Sodium Compounds; C031068:sodium perchlorate; D010751:Phosphopyruvate Hydratase