A tyrosine-tryptophan dyad and radical-based charge transfer in a ribonucleotide reductase-inspired maquette.

Zhanjun Guo,Miranda McDaniel,Gianluigi Veglia,Bridgette A. Barry,Tyler G. McCaslin,Cynthia V. Pagba,Fernando Porcelli,Jiby Yohannan

doi:10.1038/ncomms10010

Abstract

In class 1a ribonucleotide reductase (RNR), a substrate-based radical is generated in the α2 subunit by long-distance electron transfer involving an essential tyrosyl radical (Y122O·) in the β2 subunit. The conserved W48 β2 is ∼10 Å from Y122OH; mutations at W48 inactivate RNR. Here, we design a beta hairpin peptide, which contains such an interacting tyrosine–tryptophan dyad. The NMR structure of the peptide establishes that there is no direct hydrogen bond between the phenol and the indole rings. However, electronic coupling between the tyrosine and tryptophan occurs in the peptide. In addition, downshifted ultraviolet resonance Raman (UVRR) frequencies are observed for the radical state, reproducing spectral downshifts observed for β2. The frequency downshifts of the ring and CO bands are consistent with charge transfer from YO· to W or another residue. Such a charge transfer mechanism implies a role for the β2 Y-W dyad in electron transfer.

Highlights

In class 1a ribonucleotide reductase (RNR), a substrate-based radical is generated in the a2 subunit by long-distance electron transfer involving an essential tyrosyl radical (Y122O) in the b2 subunit
Peptide M is a variant of b-hairpin, Peptide A (Fig. 1c,d), in which H14 is replaced with W (Fig. 1e,f)
Long range supported by seven inter-residue contacts between the aromatics, including ROEs between the e protons of Y5 and CH2b and e of W14. These NMR experiments show that the tyrosine and tryptophan are B6 Å apart and exhibit a pi-stacked, staggered interaction, which is reminiscent of the orientation observed in the b2 dyad

Summary

Introduction

In class 1a ribonucleotide reductase (RNR), a substrate-based radical is generated in the a2 subunit by long-distance electron transfer involving an essential tyrosyl radical (Y122O) in the b2 subunit. There is no high-resolution structure of Y122O-diferric b2, in structures of the met state (Y122OHdiferric state)[7], Y122OH is buried in a hydrophobic environment and is B10 Å (Fig. 1a) from a surface-exposed tryptophan, b2 W48 This tryptophan is highly conserved in class 1a b2 sequences[8]. Mutations at W48 in E. coli b2 or at the homologous position, W103 in mouse b2 (Fig. 1b)[9], inactivate RNR8,10–12 This residue has been shown to participate in cofactor assembly[10]. Peptide M (Fig. 1e) contains a single tyrosine and a single tryptophan, which form a dyad (Y5-W14), as established here by NMR, ultraviolet absorption and circular dichroism (CD) spectropolarography. There were earlier studies of (for example, see refs 19,20) singlet states in model tyrosine–tryptophan peptides, we describe the

Methods

Results

Conclusion