Facilitating the structural characterisation of non-canonical amino acids in biomolecular NMR

Sarah Kuschert; Martin Stroet; Yanni Ka-Yan Chin; Anne Claire Conibear; Xinying Jia; Thomas Lee; Christian Reinhard Otto Bartling; Kristian Strømgaard; Peter Güntert; Karl Johan Rosengren; Alan Edward Mark; Mehdi Mobli

doi:10.5194/mr-4-57-2023

Facilitating the structural characterisation of non-canonical amino acids in biomolecular NMR

Magn Reson (Gott). 2023 Feb 24;4(1):57-72. doi: 10.5194/mr-4-57-2023. eCollection 2023.

Affiliations

¹ Centre for Advanced Imaging, Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, Brisbane, QLD 4072, Australia.
² School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD 4072, Australia.
³ Institute of Applied Synthetic Chemistry, Technische Universität Wien, Getreidemarkt 9/163, Wien 1060, Vienna, Austria.
⁴ Department of Drug Design and Pharmacology, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark.
⁵ Laboratory of Physical Chemistry, ETH Zürich, 8093 Zurich, Switzerland.
⁶ Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt, 60438 Frankfurt am Main, Germany.
⁷ Department of Chemistry, Tokyo Metropolitan University, Hachiōji, Tokyo 192-0397, Japan.
⁸ School of Biomedical Sciences, The University of Queensland, Brisbane, QLD 4072, Australia.

Abstract

Peptides and proteins containing non-canonical amino acids (ncAAs) are a large and important class of biopolymers. They include non-ribosomally synthesised peptides, post-translationally modified proteins, expressed or synthesised proteins containing unnatural amino acids, and peptides and proteins that are chemically modified. Here, we describe a general procedure for generating atomic descriptions required to incorporate ncAAs within popular NMR structure determination software such as CYANA, CNS, Xplor-NIH and ARIA. This procedure is made publicly available via the existing Automated Topology Builder (ATB) server (https://atb.uq.edu.au, last access: 17 February 2023) with all submitted ncAAs stored in a dedicated database. The described procedure also includes a general method for linking of side chains of amino acids from CYANA templates. To ensure compatibility with other systems, atom names comply with IUPAC guidelines. In addition to describing the workflow, 3D models of complex natural products generated by CYANA are presented, including vancomycin. In order to demonstrate the manner in which the templates for ncAAs generated by the ATB can be used in practice, we use a combination of CYANA and CNS to solve the structure of a synthetic peptide designed to disrupt Alzheimer-related protein-protein interactions. Automating the generation of structural templates for ncAAs will extend the utility of NMR spectroscopy to studies of more complex biomolecules, with applications in the rapidly growing fields of synthetic biology and chemical biology. The procedures we outline can also be used to standardise the creation of structural templates for any amino acid and thus have the potential to impact structural biology more generally.