Work

Recombinant Expression and Chemical Amidation of Isotopically Labeled Native Melittin

Year: 2023

Type: article

Abstract: High Resolution Image Download MS PowerPoint Slide Post-translational modifications are ubiquitous in the eukaryotic proteome. However, these modifications are rarely incorporated in NMR studies of eukaryotic proteins, which are typically produced through recombinant expression in E. coli . Melittin is the primary peptide in honey bee venom. Its native C-terminal amide significantly affects its equilibrium structure and dynamics in solution and is thus a prerequisite for studying its native structure and function. Here, we present a method for producing triply isotopically labeled ( 2 H, 13 C, and 15 N) native melittin through recombinant expression followed by chemical amidation. We then show that structural models produced with AlphaFold-Multimer are in even better agreement with experimental residual dipolar couplings than the 2.0 Å resolution X-ray crystal structure for residues G3–K23.

Source: Journal of the American Chemical Society

DOI: 10.1021/jacs.2c12631

Authors: Martin D. Gelenter, Ad Bax

Institutions: National Institutes of Health, National Institute of Diabetes and Digestive and Kidney Diseases

Language: English

Topic: Advanced NMR Techniques and Applications

Subfield: Spectroscopy

Field: Chemistry

Domain: Physical Sciences

Open Access status: hybrid

Estimated APC paid: $5,000

Funder: National Institute of Diabetes and Digestive and Kidney Diseases