Manipulation of an existing crystal form unexpectedly results in interwoven packing networks with pseudo-translational symmetry.

Janice M Reimer,Harold R Powell,T Martin Schmeing,Martin N Aloise

doi:10.1107/s2059798316013504

Janice M Reimer, Harold R Powell + Show 2 more

Open Access

https://doi.org/10.1107/s2059798316013504

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Abstract

Nonribosomal peptide synthetases (NRPSs) are multimodular enzymes that synthesize a myriad of diverse molecules. Tailoring domains have been co-opted into NRPSs to introduce further variety into nonribosomal peptide products. Linear gramicidin synthetase contains a unique formylation-tailoring domain in its initiation module (F-A-PCP). The structure of the F-A di-domain has previously been determined in a crystal form which had large solvent channels and no density for the minor Asub subdomain. An attempt was made to take advantage of this packing by removing the Asub subdomain from the construct (F-AΔsub) in order to produce a crystal that could accommodate the PCP domain. In the resulting crystal the original packing network was still present, but a second network with the same packing and almost no contact with the original network took the place of the solvent channels and changed the space group of the crystal.

Highlights

Nonribosomal peptides (NRPs), which are small molecules that are produced by nonribosomal peptide synthetases (NRPSs), cover an enormous expanse of chemical space and have uses ranging from green chemicals to important antibiotics (Walsh, 2004)
The large diversity in NRPs comes from the ability of NRPSs to use >500 monomers as substrates and to co-synthetically introduce chemical modifications into these substrates
Aimed to generate structural insight into how LgrA incorporates an F domain into its initiation module and what adaptations the various domains of the NRPS have to undergo to allow a tailoring domain to function within its system. This information could guide future bioengineering endeavours focused on incorporating formylation domains into noncognate NRPS systems

Summary

Introduction

Nonribosomal peptides (NRPs), which are small molecules that are produced by nonribosomal peptide synthetases (NRPSs), cover an enormous expanse of chemical space and have uses ranging from green chemicals to important antibiotics (Walsh, 2004). The formylation (F) domain in the F-A-PCP initiation module of the linear gramicidin synthetase (LgrA–E) from Brevibacillus parabrevis has been shown to N-formylate the first amino acid, valine, in linear gramicidin (Kessler et al, 2004). This formylation event allows elongation to continue and is required for the biological activity of linear gramicidin (Schoenafinger et al, 2006). Aimed to generate structural insight into how LgrA incorporates an F domain into its initiation module and what adaptations the various domains of the NRPS have to undergo to allow a tailoring domain to function within its system This information could guide future bioengineering endeavours focused on incorporating formylation domains into noncognate NRPS systems. The removal of the Asub subdomain resulted in a crystal featuring unexpected packing, with a second, identical but largely independent, packing network interwoven with the original network

Macromolecule production

Crystallization

Data collection and processing

Structure solution and refinement

Results and discussion