Zn(2+) chelation by serum albumin improves hexameric Zn(2+)-insulin dissociation into monomers after exocytosis

β-cells release hexameric Zn(2+)-insulin into the extracellular space, but monomeric Zn(2+)-free insulin appears to be the only biologically active form. The mechanisms implicated in dissociation of the hexamer remain unclear, but they seem to be Zn(2+) concentration-dependent. In this study, we investigate the influence of albumin binding to Zn(2+) on Zn(2+)-insulin dissociation into Zn(2+)-free insulin and its physiological, methodological and therapeutic relevance. Glucose and K(+)-induced insulin release were analyzed in isolated mouse islets by static incubation and perifusion experiments in the presence and absence of albumin and Zn(2+) chelators. Insulin tolerance tests were performed in rats using different insulin solutions with and without Zn(2+) and/or albumin. Albumin-free buffer does not alter quantification by RIA of Zn(2+)-free insulin but strongly affects RIA measurements of Zn(2+)-insulin. In contrast, accurate determination of Zn(2+)-insulin was obtained only when bovine serum albumin or Zn(2+) chelators were present in the assay buffer solution. Albumin and Zn(2+) chelators do not modify insulin release but do affect insulin determination. Preincubation with albumin or Zn(2+) chelators promotes the conversion of “slow” Zn(2+)-insulin into “fast” insulin. Consequently, insulin diffusion from large islets is ameliorated in the presence of Zn(2+) chelators. These observations support the notion that the Zn(2+)-binding properties of albumin improve the dissociation of Zn(2+)-insulin into subunits after exocytosis, which may be useful in insulin determination, insulin pharmacokinetic assays and islet transplantation.