TAILIEUCHUNG - Báo cáo khoa học: Conformational and functional analysis of the lipid binding protein Ag-NPA-1 from the parasitic nematode Ascaridia galli

Ag-NPA-1 (AgFABP), a 15 kDa lipid binding protein (LBP) fromAscari-dia galli, is a member of the nematode polyprotein allergen/antigen (NPA) family. Spectroscopic analysis shows that Ag-NPA-1 is a highly ordered, a-helical protein and that ligand binding slightly increases the ordered sec-ondary structure content. The conserved, single Trp residue (Trp17) and three Tyr residues determine the fluorescence properties of Ag-NPA-1. | iFEBS Journal Conformational and functional analysis of the lipid binding protein Ag-NPA-1 from the parasitic nematode Ascaridia galli Rositsa Jordanova1 Georgi Radoslavov1 Peter Fischer2 Eva Liebau2 Rolf D. Walter2 Ilia Bankov1 and Raina Boteva3 1 Institute of ExperimentalPathology and Parasitology Sofia Bulgaria 2 Bernhard Nocht Institute for TropicalMedicine Hamburg Germany 3 NationalCenter of Radiobiology and Radiation Protection Sofia Bulgaria Keywords NPA Trp and IAEDANS fluorescence FRET immunohistology Correspondence R. Boteva NationalCenter of Radiobiology and Radiation Protection Sofia 1756 Bulgaria Fax 359 28621059 Tel 359 28626036 210 E-mail Received 27 July 2004 revised 17 September 2004 accepted 20 September 2004 doi Ag-NPA-1 AgFABP a 15 kDa lipid binding protein LBP from Ascari-dia galli is a member of the nematode polyprotein allergen antigen NPA family. Spectroscopic analysis shows that Ag-NPA-1 is a highly ordered a-helical protein and that ligand binding slightly increases the ordered secondary structure content. The conserved single Trp residue Trp17 and three Tyr residues determine the fluorescence properties of Ag-NPA-1. Analysis of the efficiency of the energy transfer between these chromophores shows a high degree of Tyr-Trp dipole-dipole coupling. Binding of fatty acids and retinol was accompanied by enhancement of the Trp emission which allowed calculation of the affinity constants of the binary complexes. The distance between the single Trp of Ag-NPA-1 and the fluorescent fatty acid analogue 11- 5-dimethylaminonaphthalene-1- sulfo-nyl amino undecanoic acid DAUDA from the protein binding site is nm as estimated by fluorescence resonance energy transfer. A chemical modification of the Cys residues of Ag-NPA-1 Cys66 and Cys122 with the thiol reactive probes 5- 2-iodoacetyl amino ethyl amino naph-thalene-1-sulfonic acid IAEDANS and N N-dimethyl-N- iodoacetyl -N- 7-nitrobenz-2-oxa-1 .

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