- Tytuł:
- GTP-binding properties of the membrane-bound form of porcine liver annexin VI.
- Autorzy:
-
Kirilenko, Aneta
Golczak, Marcin
Pikuła, Sławomir
Bandorowicz-Pikuła, Joanna - Powiązania:
- https://bibliotekanauki.pl/articles/1044028.pdf
- Data publikacji:
- 2001
- Wydawca:
- Polskie Towarzystwo Biochemiczne
- Tematy:
-
annexin VI
GTP-binding domain
TNP-GTP
circular dichroism
GTP binding - Opis:
- Annexin VI (AnxVI) of molecular mass 68-70 kDa belongs to a multigenic family of ubiquitous Ca2+ - and phospholipid-binding proteins. In this report, we describe the GTP-binding properties of porcine liver AnxVI, determined with a fluorescent GTP analogue, 2'-(or 3')-O-(2,4,6-trinitrophenyl)guanosine 5'-triphosphate (TNP-GTP). The optimal binding of TNP-GTP to AnxVI was observed in the presence of Ca2+ and asolectin liposomes, as evidenced by a 5.5-fold increase of TNP-GTP fluorescence and a concomitant blue shift (by 17 nm) of its maximal emission wavelength. Titration of AnxVI with TNP-GTP resulted in the determination of the dissociation constant (Kd) and binding stoichiometry that amounted to 1.3 μM and 1:1 TNP-GTP/AnxVI, mole/mole, respectively. In addition, the intrinsic fluorescence of the membrane-bound form of AnxVI was quenched by TNP-GTP and this was accompanied by fluorescence resonance energy transfer (FRET) from AnxVI Trp residues to TNP-GTP. This indicates that the GTP-binding site within the AnxVI molecule is probably located in the vicinity of a Trp-containing domain of the protein. By controlled proteolysis of human recombinant AnxVI, followed by purification of the proteolytic fragments by affinity chromatography on GTP-agarose, we isolated a 35 kDa fragment corresponding to the N-terminal half of AnxVI containing Trp192. On the basis of these results, we suggest that AnxVI is a GTP-binding protein and the binding of the nucleotide may have a regulatory impact on the interaction of annexin with membranes, e.g. formation of ion channels by the protein.
- Źródło:
-
Acta Biochimica Polonica; 2001, 48, 4; 851-865
0001-527X - Pojawia się w:
- Acta Biochimica Polonica
- Dostawca treści:
- Biblioteka Nauki
Artykuł