Molecular modeling and deletion mutagenesis implicate the nuclear translocation sequence in structural integrity of fibroblast growth factor-1.
The sequence NYKKPKL in the NH2 terminus of fibroblast growth factor (FGF)-1 has been proposed to affect the long term activities of FGF-1 through its function as a nuclear translocation signal or its role in stabilization of the structure required to sustain binding and activation of the transmembrane receptor kinase. A dynamic molecular model of FGF-1 docked into a duplex of the FGF receptor ectodomain and a hexadecameric heparin chain suggests that the NYKKPKL sequence does not directly interact with heparin or the receptor, but rather the lysine-leucine residues within the sequence indirectly stabilize a major receptor-binding domain. Concurrent with a marked increase in dependence on exogenous heparin for optimal activity, sequential deletion of residues in the NYKKPKL sequence in FGF-1 resulted in a progressive loss of thermal stability, resistance to protease, mitogenic activity, and affinity for the transmembrane receptor. The largest change resulted from deletion of the entire sequence through the lysine-leucine residues. In the presence of sufficiently high concentrations of heparin, the deletion mutants exhibited mitogenic activity equal to wild-type FGF-1. The results confirm that a primary role of the NYKKPKL sequence domain is to maintain the structural integrity of FGF-1 required for optimal binding to and activation of the heparan sulfate-transmembrane receptor complex.
Center for Cancer Biology and Nutrition, Albert B. Alkek Institute of Biosciences and Technology, Department of Biochemistry and Biophysics, Texas A & M University, Houston, Texas 77030-3303, USA.
Y
Y
Luo
J L
JL
Gabriel
F
F
Wang
X
X
Zhan
T
T
Maciag
M
M
Kan
W L
WL
McKeehan
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D002467:Cell Nucleus / Q000378:metabolism*; D005346:Fibroblast Growth Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008934:Mitogens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion
D008565:Membrane Proteins; D008934:Mitogens; D011994:Recombinant Proteins; D005346:Fibroblast Growth Factors