QM/MM Investigation for Protonation States in a Bilin Reductase PcyA‐Biliverdin IXɑ Complex

• Published in: Chemphyschem 2018-08, Vol.19 (15), p.1809-1813
• Main Authors: Iijima, Eri, Gleeson, M. Paul, Unno, Masaki, Mori, Seiji
• Format: Article
• Language: English
• Subjects: Amino acids; enzyme; Hydronium ions; Neutron diffraction; Neutrons; ONIOM Calculations; oxidoreductase; PcyA; Protonation; protonation state; Quantum mechanics; Substrates
• ISSN: 1439-4235; 1439-7641
• DOI: 10.1002/cphc.201800031

Herein we report quantum mechanical/molecular mechanical (QM/MM) studies to investigate the most probable protonation states of active site amino acids and bound substrate based on a recently reported neutron diffraction structure of phycocyanobilin:ferredoxin oxidoreductase (PcyA) by Unno et al. This structure was considered to be bound in its initial state of biliverdin IXɑ (BV), which has the C‐pyrrole ring in the deprotonated state. The protonation state of BV suggested by neutron and spectroscopic studies is a stable, two‐electron reduced complex with a bound hydronium ion. Several ambiguities in the neutron structure were observed which prompted a further theoretical analysis of the structure. This structural investigation provides new understanding of the PcyA and BV protonation states not previously reported in the literature. Our calculations suggest that the hydronium ion (H3O+) is energetically unfavorable, preferentially protonating the neighboring His88 residue and that the C‐ring of BV is not protonated. The location where protons are: QM/MM investigations are reported for most probable protonation states of active site amino acid residues and the substrate (biliverdin IXɑ) of phycocyanobilin:ferredoxin oxidoreductase (PcyA). The calculations show the absence of the proton on C‐pyrrole ring of the substrate. Several experimental ambiguities of the protonation states are discussed.