Zinc enhances adiponectin oligomerization to octadecamers but decreases the rate of disulfide bond formation

Abstract

Adiponectin, a hormone secreted from adipocytes, has been shown to protect against development of insulin resistance, ischemia-reperfusion injury, and inflammation. Adiponectin assembles into multiple oligomeric isoforms: trimers, hexamers and several higher molecular weight (HMW) species. Of these, the HMW species are selectively decreased during the onset of type 2 diabetes. Despite the critical role of HMW adiponectin in insulin responsiveness, its assembly process is poorly understood. In this report, we investigated the role of divalent cations in adiponectin assembly. Purified adiponectin 18mers, the largest HMW species, did not collapse to smaller oligomers after treatment with high concentrations of EDTA. However, treatment with EDTA or another chelator DTPA inhibited the oligomerization of 18mers from trimers in vitro. Zn(2+) specifically increased the formation of 18mers when compared with Cu(2+), Mg(2+), and Ca(2+). Distribution of adiponectin oligomers secreted from zinc chelator TPEN-treated rat adipocytes skewed toward increased proportions of hexamers and trimers. While we observed presence of zinc in adiponectin purified from calf serum, the role of zinc in disulfide bonding between oligomers was examined because the process is critical for 18mer assembly. Surprisingly, Zn(2+) inhibited disulfide bond formation early in the oligomerization process. We hypothesize that initial decreases in disulfide formation rates could allow adiponectin subunits to associate before becoming locked in fully oxidized conformations incapable of further oligomerization. These data demonstrate that zinc stimulates oligomerization of HMW adiponectin and possibly other disulfide-dependent protein assembly processes.

Alteration of local adipose tissue trace element homeostasis as a possible mechanism of obesity-related insulin resistance

“…This indication is consistent with a finding that zinc deficiency augments leptin production in mice supplemented with high fat diet in comparison to the zinc-adequate high-fat fed animals [75]. Moreover, it has been noted that in primary rat adipocytes zinc enhances oligomerization of high-molecular weight adiponectin [78] being more metabolically active in comparison to low-molecular weight adiponectin [79].…”

Zinc-induced oligomerization of zinc α2 glycoprotein reveals multiple fatty acid-binding sites

“…The integrations of Figure 3F showed that the oligomers increased with decrease in ZAG monomer in the case of zinc, but not with barium. These protein-zinc oligomers have been seen previously; the human complement regulator factor H, complement protein C3 [30], and adiponectin also form oligomers in the presence of zinc [31], while zinc induces dimerization of the class II MHC molecule HLA-DR1 in a complex with a hemagglutinin peptide [32].…”

Extracellular conversion of adiponectin hexamers into trimers

“…The reason behind the apparent discrepancy is most probably the differential staining of the oligomers by the Coomassie Brilliant Blue G250 dye. We have observed previously that the staining intensity of adiponectin trimers appeared to be lower than that of the 18mer HMW species in Coomassie dyes (Figure 1 of [35] and Figures 2 and 3 of [36]). Adiponectin is highly negatively charged at neutral pH (calculated pI approximately 5.4) and binds strongly to anion exchange columns.…”

“…Approximately 95% of purified adiponectin consists of 18mers with the balance being hexamers. Hexamers were derived from purified 18mers by adjusting the pH to 4 using glycine and re-neutralized to pH 7 using Hepes or PBS at pH 7.4 as previously described [35,36]. …”