Resolving near-ultraviolet circular dichroism spectra of single trp mutants in tear lipocalin.
Near-ultraviolet circular dichroism (near-UV CD) spectra of tryptophan residues in proteins are complicated because the line shapes are derived from the overlap of both the 1L(a) and the 1L(b) electronic bands that vary independently. Contributing to this complexity, tryptophan near-UV CD spectra differ in the relative amplitude of the 0-0 vibronic band compared to the rest of the 1L(b) spectrum, an inherent feature that may result in poor fitting. To resolve this problem, a computer program that incorporated the separation of the 0-0 transition of 1L(b) component from the rest of the 1L(b) was written in LabVIEW and its amplitude was allowed to vary independently. This method showed dramatically improved fitting of 1L(a) and 1L(b) components in the near-UV CD tryptophan spectra in tear lipocalin mutants featuring low intensity of the 0-0 1L(b) component. Side chain dynamic characteristics (mobility and accessibility to the solvent) identified from different spectroscopic techniques were related to differences in Trp near-UV CD spectra. This method is broadly applicable to different types of Trp near-UV CD spectra.
Gasymov
Oktay K
OK
Department of Pathology and Department of Ophthalmology, UCLA School of Medicine, Los Angeles, CA 90095, USA.
Abduragimov
Adil R
AR
Yusifov
Taleh N
TN
Glasgow
Ben J
BJ
D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics*; D002942:Circular Dichroism / Q000379:methods*; D054835:Lipocalin 1; D009154:Mutation / Q000235:genetics*; D011487:Protein Conformation; D015203:Reproducibility of Results; D014364:Tryptophan / Q000235:genetics*
D002352:Carrier Proteins; D054835:Lipocalin 1; D014364:Tryptophan