Department of Chemistry, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, 464-8602, Japan.
RIKEN SPring-8 Centre, 1-1-1 Kouto, Sayo, Hyogo, 679-5148, Japan.
Angew Chem Int Ed Engl. 2020 May 4;59(19):7611-7618. doi: 10.1002/anie.201913407. Epub 2020 Mar 11.
Despite CYP102A1 (P450BM3) representing one of the most extensively researched metalloenzymes, crystallisation of its haem domain upon modification can be a challenge. Crystal structures are indispensable for the efficient structure-based design of P450BM3 as a biocatalyst. The abietane diterpenoid derivative N-abietoyl-l-tryptophan (AbiATrp) is an outstanding crystallisation accelerator for the wild-type P450BM3 haem domain, with visible crystals forming within 2 hours and diffracting to a near-atomic resolution of 1.22 Å. Using these crystals as seeds in a cross-microseeding approach, an assortment of P450BM3 haem domain crystal structures, containing previously uncrystallisable decoy molecules and diverse artificial metalloporphyrins binding various ligand molecules, as well as heavily tagged haem-domain variants, could be determined. Some of the structures reported herein could be used as models of different stages of the P450BM3 catalytic cycle.
尽管 CYP102A1（P450BM3）是研究最多的金属酶之一，但对其血红素结构域进行修饰后的结晶仍然是一个挑战。对于 P450BM3 作为生物催化剂的高效基于结构的设计，晶体结构是必不可少的。落叶松二萜衍生物 N-阿魏酰-L-色氨酸（AbiATrp）是野生型 P450BM3 血红素结构域出色的结晶促进剂，在 2 小时内可见晶体形成，并衍射至近原子分辨率为 1.22 Å。使用这些晶体作为交叉微接种方法中的种子，可以确定各种 P450BM3 血红素结构域晶体结构，其中包含以前无法结晶的诱饵分子和各种结合不同配体分子的人工金属卟啉，以及标记严重的血红素结构域变体。本文报道的一些结构可作为 P450BM3 催化循环不同阶段的模型。
