Automatic structure-based NMR methyl resonance assignment in large proteins

Iva Pritišanac, Julia M. Würz, T. Reid Alderson, and Peter Güntert

Apr 09, 2019

Received Date: 1st April 19

Isotope-labeled methyl groups provide NMR probes that can be observed in very large, otherwise deuterated systems and enable investigations of protein structure, dynamics and mechanisms. However, the assignment of resonances to specific methyls in the protein is expensive and time-consuming, which limits the use of methyl-based NMR for large proteins. To resolve this bottleneck, methyl assignment methods have been developed. However, these remain limited regarding complete automation, computational feasibility, and/or the extent and accuracy of the assignments. Here, we present the automated MethylFLYA method for the assignment of methyl groups that is based on methyl-methyl nuclear Overhauser effect spectroscopy (NOESY) peak lists. MethylFLYA was applied to five proteins (28­–358 kDa) comprising a total of 708 isotope-labeled methyl groups, of which 674 had manually determined 1H/13C reference assignments and 614 showed cross peaks in the available NOESY peak lists. MethylFLYA confidently assigned 488 methyl groups, i.e. 79% of those with NOESY data. Of these, 460 agreed with the reference, 5 were different, and 23 concerned methyls without reference assignment. For high-quality NOESY spectra, automatic NOESY peak picking followed by resonance assignment with MethylFLYA can yield results comparable to those obtained from manually prepared peak lists, indicating the feasibility of unbiased, fully automatic methyl resonance assignment starting directly from the NMR spectra. Overall, MethylFLYA assigns significantly more methyl groups than other algorithms, has an average error rate of 1%, modest runtimes of 0.4–1.2 h, and flexibility to handle arbitrary isotope labeling patterns and include data from other types of NMR spectra.

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This is an abstract of a preprint hosted on an independent third party site. It has not been peer reviewed but is currently under consideration at Nature Communications.

Nature Communications

Nature Research, Springer Nature