FKBP22 of a psychrophilic bacterium, Shewanella sp. SIB1 (SIB1 FKBP22), is a member of peptidyl-prolyl cis-trans isomerase (PPIase) and consists of N- and C-domains responsible for chaperone-like and PPIase catalytic activities, respectively. The chaperone-like activity of SIB1 FKBP22 was previously evidenced by its ability to prevent dithiothreitol (DTT)-induced insulin aggregation. Nevertheless, the mechanism by which this protein inhibits the aggregation remains unclear. To address this, the binding affinity of SIB1 FKBP22 to the native or reduced states of insulin was examined using surface plasmon resonance (SPR). The native and reduced states refer to insulin in the absence or DTT presence, respectively. The SPR sensorgram showed that SIB1 FKBP22 binds specifically to the reduced state of insulin, with a K_D value of 37.31 ± 3.20 μM. This binding was facilitated by the N-domain, as indicated by the comparable K_D values of the N-domain and SIB1 FKBP22. Meanwhile, the reduced state of insulin was found to have no affinity towards the C-domain. The K_D value of SIB1 FKBP22 was slightly decreased by NaCl but was not severely affected by FK506, a specific FKBP inhibitor. Similarly, the prevention of DTT-induced aggregation by SIB1 FKBP22 was also modulated by the N-domain and was not affected by FK506. Further, the reduced and native states of insulin had no effect on the catalytic efficiency (k_cat/K_M) of SIB1 FKBP22 towards a peptide substrate. Nevertheless, the reduced state of insulin slightly reduced the catalytic efficiency towards refolding RNase T1, at up to 1.5-fold lower than in the absence of insulin. These results suggested that the binding event was mainly facilitated by hydrophobic interaction and was independent from its PPIase activity. Altogether, a possible mechanism by which SIB1 FKBP22 prevents DTT-induced insulin aggregation was proposed.

嗜冷细菌Shewanella sp. 的FKBP22 。SIB1 (SIB1 FKBP22)，是肽基-脯氨酰顺-反的成员异构酶 (PPIase)，由分别负责分子伴侣样和 PPIase 催化活性的 N 域和 C 域组成。SIB1 FKBP22 的分子伴侣样活性先前已通过其防止二硫苏糖醇 (DTT) 诱导的胰岛素聚集的能力得到证实。然而，该蛋白质抑制聚集的机制仍不清楚。为了解决这个问题，使用表面等离子体共振 (SPR) 检查了 SIB1 FKBP22 对天然或还原状态的胰岛素的结合亲和力。天然和还原状态分别是指不存在或存在 DTT 的胰岛素。SPR 传感图显示 SIB1 FKBP22 与胰岛素的还原状态特异性结合，K _D值 37.31 ± 3.20 μM。N 域促进了这种结合，如N 域和 SIB1 FKBP22的可比K _D值所示。同时，发现胰岛素的还原状态对 C 域没有亲和力。SIB1 FKBP22的K _D值被 NaCl 轻微降低，但不受 FK506（一种特定的 FKBP 抑制剂）的严重影响。同样，SIB1 FKBP22 对 DTT 诱导的聚集的预防也受 N 结构域的调节，不受 FK506 的影响。此外，胰岛素的还原状态和天然状态对催化效率没有影响（k _cat / K _M) SIB1 FKBP22 朝向肽底物。然而，胰岛素的还原状态略微降低了对重新折叠 RNase T1 的催化效率，比没有胰岛素的情况下低 1.5 倍。这些结果表明结合事件主要由疏水相互作用促进并且独立于其 PPIase 活性。总之，提出了 SIB1 FKBP22 阻止 DTT 诱导的胰岛素聚集的可能机制。