ISSN:
0730-2312
Schlagwort(e):
FGF
;
receptors
;
internalization
;
photoactivable cross-linker
;
heparan sulfate proteoglycans
;
Life and Medical Sciences
;
Cell & Developmental Biology
Quelle:
Wiley InterScience Backfile Collection 1832-2000
Thema:
Biologie
,
Chemie und Pharmazie
,
Medizin
Notizen:
The internalization of basic fibroblast growth factor (FGF-2) was studied in Chinese hamster lung fibroblasts (CCL39). Recombinant FGF-2 was derivatized with a photoactivable agent, N-hydroxysuccinimidyl-4-azido-benzoate (HSAB), iodinated, and used to visualize intracellular FGF-2-affinity-labeled molecules after internalization at 37°C. Iodinated HSAB-FGF-2 maintained the properties of natural FGF-2 such as affinity for heparin, binding to Bek and Flg receptors, interaction with high- and low-affinity binding sites, and reinitiating of DNA synthesis in CCL39 cells. Affinity-labeling experiments at 4°C with 125I-HSAB-FGF-2 led to the detection of several FGF-cell surface complexes with apparent molecular mass of 80, 100, 125, 150, 170-180, 220, 260, and about 320 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), whereas two specific bands at 80 and 130-160 kDa were obtained using the homobifunctional cross-linking reagent, disuccinimidyl suberate. When the cells, preincubated with 125I-HSAB-FGF-2 at 4°C and then washed, were shifted to 37°C, irradiation of the internalized labeled FGF-2 led to detection of a similar but fainted profile with one major specific band at 80 kDa. Heparitinase II treatment of the cells reduced binding of 125I-HSAB-FGF-2 to its cell surface sites by 80% and internalization by 55%, indicating the involvement of heparan sulfate proteoglycans in these processes. Among the heparitinase-sensitive bands was the 80-kDa complex. © 1996 Wiley-Liss, Inc.
Zusätzliches Material:
4 Ill.
Materialart:
Digitale Medien
