Linked Life Data

8672513

Source:http://linkedlifedata.com/resource/pubmed/id/8672513

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
24
pubmed:dateCreated
1996-8-15
pubmed:abstractText
Spectroscopic studies have been performed to characterize the solution structure of the V66W mutant of Staphylococcal nuclease and the corresponding 1-136 fragment, referred to as V66W'. Whereas wild-type nuclease has a single tryptophan residue at position 140, the V66W mutant has a second tryptophan residue at position 66, which is the only such residue in V66W'. Steady-state and time-resolved fluorescence studies show Trp-66 in V66W' to have a blue emission, a relatively large fluorescence quantum yield, a long lifetime, a significant degree of protection from solute quenchers, and to depolarize with a relatively long rotational correlation time. These results characterize Trp-66 in V66W' as being a buried residue, which indicates that this fragment retains some global structure. Circular dichroism (CD) data are consistent with the fragment having lost most of the alpha-helical content of the wild type, while retaining beta-sheet structure. The CD spectrum in the aromatic region also suggests that Trp-66 in the fragment experiences an asymmetric environment, which is not identical to that in the full length mutant, V66W. In addition, optical detection of triplet state magnetic resonance (ODMR) spectroscopy can clearly resolve the tryptophan residues and demonstrates differences between the local environment of Trp-66 in V66W and in V66W', as well as small differences in the Trp-140 environment in wild type and in V66W. Guanidine-HCl induced and thermally induced unfolding studies were performed by simultaneously acquiring CD and fluorescence data as a function of the perturbation and then performing a global analysis of such multiple data sets in terms of two-state and three-state unfolding models. Whereas data for wild-type nuclease and the V66W' fragment are well characterized by a two-state unfolding model, data for the V66W mutant are better characterized by a three-state process. That is, both the denaturant- and temperature-induced unfolding of V66W involves the significant population of an equilibrium unfolding intermediate. Our global analyses yield thermodynamic parameters for the unfolding transitions, and we show that the data for V66W can be described by a constrained three-state model in which the transition of the intermediate to the fully unfolded state is fixed to have the same thermodynamic parameters that describe the unfolding of the V66W' fragment.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
18
pubmed:volume
35
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
8084-94
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed-meshheading:8672513-Calorimetry, pubmed-meshheading:8672513-Circular Dichroism, pubmed-meshheading:8672513-Escherichia coli, pubmed-meshheading:8672513-Guanidine, pubmed-meshheading:8672513-Guanidines, pubmed-meshheading:8672513-Mathematics, pubmed-meshheading:8672513-Micrococcal Nuclease, pubmed-meshheading:8672513-Models, Theoretical, pubmed-meshheading:8672513-Mutagenesis, Site-Directed, pubmed-meshheading:8672513-Peptide Fragments, pubmed-meshheading:8672513-Point Mutation, pubmed-meshheading:8672513-Protein Conformation, pubmed-meshheading:8672513-Protein Denaturation, pubmed-meshheading:8672513-Recombinant Proteins, pubmed-meshheading:8672513-Spectrometry, Fluorescence, pubmed-meshheading:8672513-Staphylococcus, pubmed-meshheading:8672513-Thermodynamics, pubmed-meshheading:8672513-Tryptophan
pubmed:year
1996
pubmed:articleTitle
Thermodynamics of the unfolding and spectroscopic properties of the V66W mutant of Staphylococcal nuclease and its 1-136 fragment.
pubmed:affiliation
Department of Chemistry, University of Mississippi, University 38677, USA.
pubmed:publicationType
Journal Article, Comparative Study, Research Support, U.S. Gov't, P.H.S., Research Support, U.S. Gov't, Non-P.H.S.