College of Pharmacy, University of New Mexico, Albuquerque, NM, USA.
Department of Chemistry and Chemical Biology, University of New Mexico, Albuquerque, NM, USA.
FEBS Lett. 2018 Jul;592(14):2425-2431. doi: 10.1002/1873-3468.13158. Epub 2018 Jun 29.
The interface between calmodulin (CaM) and the NO synthase (NOS) heme domain is the least characterized interprotein interface that the NOS isoforms must traverse through during catalysis. Our previous molecular dynamics simulations predicted a salt bridge between K497 in human inducible NOS (iNOS) heme domain and D118(CaM). Herein, the FMN - heme interdomain electron transfer (IET) rate was found to be notably decreased by charge-reversal mutation, while the IET in the iNOS K497D mutant is significantly restored by the CaM D118K mutation. The results of wild-type protein with added synthetic peptides further demonstrate the critical nature of K497 relative to the rest of the peptide sequence in modulating the IET. These data provide definitive evidence supporting the regulatory role of the isoform-specific K497 residue.
钙调蛋白（CaM）与一氧化氮合酶（NOS）血红素结构域之间的界面是NOS 同工酶在催化过程中必须穿越的最具特征性的蛋白间界面。我们之前的分子动力学模拟预测，人类诱导型 NOS（iNOS）血红素结构域中的 K497 与 CaM 的 D118 之间存在盐桥。在此，发现带正电荷的突变显著降低了 FMN-血红素结构域间电子转移（IET）的速率，而 iNOS K497D 突变体中的 IET 则通过 CaM D118K 突变显著恢复。加入合成肽的野生型蛋白的结果进一步证明了 K497 相对于肽序列其余部分在调节 IET 方面的关键作用。这些数据提供了明确的证据，支持同工酶特异性 K497 残基的调节作用。
