To be filled out by faculty nominator, can be edited by student:
Dr. Ramakrishna Guda
Denaturation of the N-terminal domains 5-6 of Wilson Disease protein has been monitored through its mutant WLN5-6Cys51 by circular dichroism (CD), one and two-photon fluorescence spectroscopy after binding it to the thiolreactive dye 7-diethylamino-3-(4‘-maleimidylphenyl) -4-methylcoumarin (CPM). Far-UV CD measurements have shown that the mutations and the binding of the dye did not affect the secondary structure of the protein as the unfolding transition remained constant. The fluorescence maximum of the dye changed with the denaturant concentration and followed the trend obtained from CD and shown that one dye labeled protein can be used to monitor the unfolding of protein. Interestingly, the two-photon absorption (TPA) cross-section (ratio of twophoton to one-photon fluorescence intensity) of the dye increased 40% when it is unfolded and no such change was observed for the control dye. The variations in TPA cross-section are attributed to the changes in local electric fields of the protein as confirmed from femtosecond fluorescence anisotropy and fluorescence lifetime. Present results show an evidence that the TPA cross-sections can be used to monitor the local electric fields in proteins and thereby image the folded and unfolded forms of protein or metal binding to the proteins.
WMU ScholarWorks Citation
Alja'afreh, Ibtesam Y.; Guda, Ramakrishna; and Huffman, David, "Multiphoton Fluorescence Spectroscopy to Monitor the Folding/Unfolding of the N-terminal Domains 5-6 of Wilson Disease Protein" (2012). Research and Creative Activities Poster Day. 68.