NMR studies on the equilibrium unfolding of ketosteroid isomerase by urea.
Multidimensional NMR was employed to investigate the structural changes in the urea-induced equilibrium unfolding of the dimeric ketosteroid isomerase (KSI) from Pseudomonas putida biotype B. Sequence specific backbone assignments for the native KSI and the protein with 3.5 M urea were carried out using various 3D NMR experiments. Hydrogen exchange measurements indicated that the secondary structures of KSI were not affected significantly by urea up to 3.5 M. However, the chemical shift analysis of 1H-(15)N HSQC spectra at various urea concentrations revealed that the residues in the dimeric interface region, particularly around the beta5-strand, were significantly perturbed by urea at low concentrations, while the line-width analysis indicated the possibility of conformational exchange at the interface region around the beta6-strand. The results thus suggest that the interface region primarily around the beta5- and beta6-strands could play an important role as the starting positions in the unfolding process of KSI.
Lee
Hyeong Ju
HJ
Department of Chemistry; and Division of Molecular Life Sciences, Pohang University of Science and Technology, Pohang, Republic of Korea, 790-784.
Jang
Do Soo
DS
Cha
Hyung Jin
HJ
Moon
Hye Seon
HS
Hong
Bee Hak
BH
Choi
Kwan Yong
KY
Lee
Hee Cheon
HC
D019281:Dimerization; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D011489:Protein Denaturation; D017510:Protein Folding; D013251:Steroid Isomerases / Q000737:chemistry*; D014508:Urea / Q000737:chemistry*
D014508:Urea; D013251:Steroid Isomerases