Glutathione and Vitamin B-12 Cooperate in Stabilization of a B-12 Trafficking Chaperone Protein

Title
Glutathione and Vitamin B-12 Cooperate in Stabilization of a B-12 Trafficking Chaperone Protein
Author(s)
김지회박지현[박지현]
Keywords
ISOTHERMAL DENATURATION; HUMAN-DISEASES; STABILITY; AFFINITY; BINDING
Issue Date
201202
Publisher
SPRINGER
Citation
PROTEIN JOURNAL, v.31, no.2, pp.158 - 165
Abstract
The protein bCblC (bCblCpro) is a bovine homolog of a human B-12 trafficking chaperone that is responsible for the processing of vitamin B-12 and its escorted delivery in intracellular B-12 metabolism. In this study, we found that bCblCpro is highly thermolabile with a T-m = 42.0 +/- 0.2 degrees C as shown for the human homolog, suggesting thermal regulation of these proteins. Binding of the reduced form of glutathione (GSH) that is a predominant cellular thiol increased the Tm of bCblCpro from 42 degrees C to similar to 45 degrees C (Delta T-m max = 3.1 +/- 0.2 degrees C and AC(50) = 2.1 +/- 0.5 mM). Binding of vitamin B-12 and its derivatives also stabilized bCblCpro increasing the Tm to a different extent and vitamin B-12 (cyanocobalamin, CNCbl) was the least efficient (Delta T-m max = 4.3 +/- 0.3 degrees C and AC(50) = 291 +/- 36 mu M). However, the stabilizing effect of CNCbl was significantly greater for GSH-bound bCblCpro (Delta T-m max = 12.8 +/- 0.6 degrees C and AC(50) = 9.3 +/- 1.6 mu M) than for GSH-free bCblCpro. In addition, the stabilizing effect of GSH was also greater for CNCbl-bound bCblCpro (Delta T-m max = 9.3 +/- 0.3 degrees C and AC(50) = 57.0 +/- 6.8 mu M). Limited proteolysis revealed that thermal stabilization of bCblCpro is derived from conformational changes of the protein induced by binding of the ligands. The results in this study indicate that GSH cooperates with vitamin B-12 in thermal stabilization of bCblCpro and is a positive regulator of the protein.
URI
http://hdl.handle.net/YU.REPOSITORY/29921http://dx.doi.org/10.1007/s10930-011-9385-2
ISSN
1572-3887
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