Hydrogen-exchange kinetics of the indole NH proton of the buried tryptophan in the constant fragment of the immunoglobulin light chain.
The constant fragment of the immunoglobulin light chain (type lambda) has two tryptophyl residues at positions 150 and 187. Trp-150 is buried in the interior, and Trp-187 lies on the surface of the molecule. The hydrogen-deuterium exchange kinetics of the indole NH proton of Trp-150 were studied at various pH values at 25 degrees C by 1H nuclear magnetic resonance. Exchange rates were approximately first order in hydroxyl ion dependence above pH 8, were relatively independent of pH between pH 7 and 8, and decreased below pH 7. On the assumption that the exchange above pH 8 proceeds through local fluctuations of the protein molecule, the exchange rates between pH 7 and 8 through global unfolding were estimated. The exchange rate constant within this pH range at 25 degrees C thus estimated was consistent with that of the global unfolding of the constant fragment under the same conditions as those reported previously [Kikuchi, H., Goto, Y., & Hamaguchi, K. (1986) Biochemistry 25, 2009-2013]. The activation energy for the exchange process at pH 7.8 was the same as that for the unfolding process by 2 M guanidine hydrochloride. The exchange rates of backbone NH protons were almost the same as that of the indole NH proton of Trp-150 at pH 7.1. These observations also indicated that the exchange between pH 7 and 8 occurs through global unfolding of the protein molecule and is rate-limited by the unfolding.(ABSTRACT TRUNCATED AT 250 WORDS)
Kawata
Y
Y
Department of Biology, Faculty of Science, Osaka University, Japan.
Goto
Y
Y
Hamaguchi
K
K
Hayashi
F
F
Kobayashi
Y
Y
Kyogoku
Y
Y
D001536:Bence Jones Protein; D003903:Deuterium; D006801:Humans; D006859:Hydrogen; D006863:Hydrogen-Ion Concentration; D007147:Immunoglobulin Light Chains; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D010206:Papain; D011487:Protein Conformation; D011489:Protein Denaturation; D011522:Protons; D014364:Tryptophan
D007147:Immunoglobulin Light Chains; D011522:Protons; D006859:Hydrogen; D014364:Tryptophan; D001536:Bence Jones Protein; D003903:Deuterium; D010206:Papain