Modification of a specific tyrosine enables tracing of the end‐to‐end distance during apomyoglobin folding

[摘要]

In order to follow the overall geometry of the apomyoglobin molecule during folding, we have converted a specific tyrosine residue into 3-nitro-tyrosine. The specificity of the modification was verified by proteolytic cleavage of the modified protein and mass spectroscopy of the resulting fragments. By measuring the energy transfer from the tryptophanyl side-chains to the modified residue the average end-to-end distance can be followed. The experiment shows that after initiation of folding the N- and C-termini are rapidly brought into proximity, possibly to a near-native distance.

[学科分类] 生物化学/生物物理

[关键词] Protein folding;Energy transfer;Nitro-tyrosine;Stopped-flow;CD;circular dichroism;AEDANS;5-(((acetylamino)ethyl)amino) naphthalene-1-sulfonic acid;apoMb;apomyoglobin;Tyr(NO2);3-nitro-tyrosine;apoMb-Y146(NO2);apoMb with Tyr146 converted to Tyr(NO2);SDS-PAGE;sodium dodecyl sulfate polyacryl-amide gel electrophoresis;TNM;tetranitromethane;HPLC;high performance liquid chromatography;MALDI-TOF;matrix assisted laser desorption ionization - time-of-flight (mass spectroscopy);ACH;α-cyano-4-hydroxy-cinnamic acid;GuHCl;guanidine-hydrochloride