It is well known that the SEA-TROSY experiment could alleviate some of the problems of resonance overlap in 15N/2H labeled proteins as it was designed to selectively map solvent exposed amide protons. However, SEA- TR...It is well known that the SEA-TROSY experiment could alleviate some of the problems of resonance overlap in 15N/2H labeled proteins as it was designed to selectively map solvent exposed amide protons. However, SEA- TROSY spectra may be contaminated with exchange-relayed NOE contributions from fast exchanged hydroxyl or amine protons and contributions from longitudinal relaxation. Also, perdeuteration of the protein sample is a pre-requisite for this experiment. In this communication, a modified version, clean SEA-TROSY, was proposed to eliminate these artifacts and to allow the experiment to be applied to protonated or partially deuterated proteins and protein complexes.展开更多
This paper describes an amide-exchange-rate-edited （AERE） NMR method that can effectively alleviate the problem of resonance overlap for proteins and peptides. This method exploits the diversity of amide proton exch...This paper describes an amide-exchange-rate-edited （AERE） NMR method that can effectively alleviate the problem of resonance overlap for proteins and peptides. This method exploits the diversity of amide proton exchange rates and consists of two complementary experiments： （1） SEA （solvent exposed amide）-type NMR experiments to map exchangeable surface residues whose amides are not involved in hydrogen bonding, and （2） presat-type NMR experiments to map solvent inaccessibly buried residues or nonexchangeable residues located in hydrogen-bonded secondary structures with properly controlled saturation transfer via amide proton exchanges with the solvent. This method separates overlapping resonances in a spectrum into two complementary spectra. The AERE-NMR method was demonstrated with a sample of ^15N/^13C/^2H（70%） labeled ribosome-inactivating protein trichosanthin of 247 residues.展开更多
基金Project supported by the grants from One Hundred Talent Program of the Chinese Academy of Sciences and Shanghai Commission of Science and Technology (No. 03JC14081).
文摘It is well known that the SEA-TROSY experiment could alleviate some of the problems of resonance overlap in 15N/2H labeled proteins as it was designed to selectively map solvent exposed amide protons. However, SEA- TROSY spectra may be contaminated with exchange-relayed NOE contributions from fast exchanged hydroxyl or amine protons and contributions from longitudinal relaxation. Also, perdeuteration of the protein sample is a pre-requisite for this experiment. In this communication, a modified version, clean SEA-TROSY, was proposed to eliminate these artifacts and to allow the experiment to be applied to protonated or partially deuterated proteins and protein complexes.
基金Project supported by the National Natural Science Foundation of China （No. 30470351）.Acknowledgement We would like to thank Prof. Zhu G. and Dr. Sze K. H. for help with this work. We also thank Mr. Cheung K. K. for help with the backbone resonance assignment of the TCS protein.
文摘This paper describes an amide-exchange-rate-edited （AERE） NMR method that can effectively alleviate the problem of resonance overlap for proteins and peptides. This method exploits the diversity of amide proton exchange rates and consists of two complementary experiments： （1） SEA （solvent exposed amide）-type NMR experiments to map exchangeable surface residues whose amides are not involved in hydrogen bonding, and （2） presat-type NMR experiments to map solvent inaccessibly buried residues or nonexchangeable residues located in hydrogen-bonded secondary structures with properly controlled saturation transfer via amide proton exchanges with the solvent. This method separates overlapping resonances in a spectrum into two complementary spectra. The AERE-NMR method was demonstrated with a sample of ^15N/^13C/^2H（70%） labeled ribosome-inactivating protein trichosanthin of 247 residues.
