Characterization of the weak calcium binding of trimeric globular adiponectin

摘要

Adiponectin is secreted from adipose tissue and functions as a protein hormone in regulating glucose metabolism and fatty acid catabolism. Adiponectin plays an important role as a novel risk factor and potential diagnostic and prognostic biomarker in cancer. Crystal structures of globular adiponectin have been resolved with three calcium-binding sites on the top of its central tunnel. However, the calcium-binding property of adiponectin remains elusive. Mouse globular adiponectin was cloned into pET11a and expressed in Escherichia coli. The folding of adiponectin was indicated by the spread of resonances in HSQC spectrum. Luminescence resonance energy transfer was used to obtain the binding constant (Kd) of Tb3+ and the inhibitor constant (Ki) of Ca2+ for globular adiponectin. The obtained calcium-binding affinity to adiponectin is relatively low (~2mM), which indicates that the high concentration of adiponectin in circulating system may function as calcium storage bank and buffer the free calcium concentration.