Pyrene excimer fluorescence of yeast alcohol dehydrogenase: a sensitive probe to investigate ligand binding and unfolding pathway of the enzyme

Photochemistry and Photobiology
Manas Kumar SantraDulal Panda

Abstract

The cysteine residues of yeast alcohol dehydrogenase (YADH) were covalently modified by N-(1-pyrenyl) maleimide (PM). A maximum of 3.4 cysteines per YADH monomer could be modified by PM. The secondary structure of PM-YADH was found to be similar to that of the native YADH using far-UV circular dichroism. The covalent modification of YADH by PM inhibited the enzymatic activity indicating that the active site of the enzyme was altered. PM-YADH displayed maximum excimer fluorescence at an incorporation ratio of 2.6 mol of PM per monomeric subunit of YADH. Nucleotide adenine dinucleotide (NAD) divalent zinc and ethanol reduced the excimer fluorescence of PM-YADH indicating that these agents induce conformational changes in the enzyme. Guanidinium hydrochloride (GdnHCl)-induced unfolding of YADH was analyzed using tryptophan fluorescence, pyrene excimer fluorescence and enzymatic activity. The unfolding of YADH was found to occur in a stepwise manner. The loss of enzymatic activity preceded the global unfolding of the protein. Further, changes in tryptophan fluorescence with increasing GdnHCl suggested that YADH was completely unfolded by 2.5 M GdnHCl. Interestingly, residual structures of YADH were detected even in the presence of ...Continue Reading

References

Mar 25, 1976·Journal of Molecular Biology·H EklundA Akeson
Jun 7, 1976·Biochemical and Biophysical Research Communications·J P Klinman, K Welsh
Dec 15, 1975·Biochemical and Biophysical Research Communications·C Veillon, A J Sytkowski
Mar 1, 1992·European Journal of Biochemistry·D Panda, B Bhattacharyya
Apr 6, 1992·FEBS Letters·L Y Cheng, L H Lek
Jan 1, 1990·Proceedings of the National Academy of Sciences of the United States of America·B L Vallee, D S Auld
Mar 1, 1964·The Biochemical Journal·E P Whitehead, B R Rabin
Jul 1, 1965·Journal of Biochemistry·T Ohta, Y Ogura
Aug 1, 1973·Proceedings of the National Academy of Sciences of the United States of America·C I BrändénA Akeson
Feb 1, 1968·Proceedings of the National Academy of Sciences of the United States of America·B L Vallee, R J Williams
Aug 1, 1996·International Journal of Biological Macromolecules·T Zhang, H M Zhou
Mar 10, 2001·Biochemistry. Biokhimii︠a︡·Y Yang, H M Zhou
Apr 19, 2003·FEMS Yeast Research·Vladimir LeskovacDraginja Pericin

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Citations

Oct 4, 2012·Free Radical Biology & Medicine·Naomi L CookMichael J Davies
Nov 8, 2018·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Yongxian FanYinchu Shen
Dec 7, 2011·Dalton Transactions : an International Journal of Inorganic Chemistry·Nicola M CoxHarry Adams
Jan 30, 2021·Journal of Asian Natural Products Research·Zhi MaXiao-Long Chen

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Methods Mentioned

BETA
HLADH
gel filtration

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