Elucidation of substrate binding interactions in a membrane transport protein by mass spectrometry
| Jazyk: | English |
|---|---|
| Přístupová URL adresa: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::cfd59f91c40704a6d86c7adde36aa384 https://europepmc.org/articles/PMC152890/ |
| Rights: | OPEN |
| Přírůstkové číslo: | edsair.doi.dedup.....cfd59f91c40704a6d86c7adde36aa384 |
| Autor: | Adam B. Weinglass, G. Verner, Kym F. Faull, Yonglin Hu, H. Ronald Kaback, Julian P. Whitelegge |
| Jazyk: | angličtina |
| Rok vydání: | 2003 |
| Předmět: |
Lactose permease
Spectrometry Mass Electrospray Ionization Monosaccharide Transport Proteins Stereochemistry Protein Conformation Electrospray ionization Biology General Biochemistry Genetics and Molecular Biology Substrate Specificity chemistry.chemical_compound Protein structure Escherichia coli Molecular Biology Integral membrane protein Chromatography High Pressure Liquid Carbodiimide General Immunology and Microbiology Symporters Permease General Neuroscience Escherichia coli Proteins Substrate (chemistry) Membrane Transport Proteins Articles Ligand (biochemistry) Peptide Fragments Carbodiimides Biochemistry chemistry Amino Acid Substitution Protein Binding |
| Popis: | Integration of biochemical and biophysical data on the lactose permease of Escherichia coli has culminated in a molecular model that predicts substrate-protein proximities which include interaction of a hydroxyl group in the galactopyranosyl ring with Glu269. In order to test this hypothesis, we studied covalent modification of carboxyl groups with carbodiimides using electrospray ionization mass spectrometry (ESI-MS) and demonstrate that substrate protects the permease against carbodiimide reactivity. Further more, a significant proportion of the decrease in carbodiimide reactivity occurs specifically in a nanopeptide containing Glu269. In contrast, carbodiimide reactivity of mutant Glu269-->Asp that exhibits lower affinity is unaffected by substrate. By monitoring the ability of different substrate analogs to protect against carbodiimide modification of Glu269, it is suggested that the C-3 OH group of the galactopyranosyl ring may play an important role in specificity, possibly by H-bonding with Glu269. The approach demonstrates that mass spectrometry can provide a powerful means of analyzing ligand interactions with integral membrane proteins. |
| Databáze: | OpenAIRE |
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