Structural basis of conserved cysteine in the fibroblast growth factor family: evidence for a vestigial half-cystine.
The 22 members of the mouse/human fibroblast growth factor (FGF) family of proteins contain a conserved cysteine residue at position 83 (numbering scheme of the 140-residue form of FGF-1). Sequence and structure information suggests that this position is a free cysteine in 16 members and participates as a half-cystine in at least 3 (and perhaps as many as 6) other members. While a structural role as a half-cystine provides a stability basis for possible selective pressure, it is less clear why this residue is conserved as a free cysteine (although free buried thiols can limit protein functional half-life). To probe the structural role of the free cysteine at position 83 in FGF-1, we constructed Ala, Ser, Thr, Val, and Ile mutations and determined their effects on structure and stability. These results show that position 83 in FGF-1 is thermodynamically optimized to accept a free cysteine. A second cysteine mutation was introduced into wild-type FGF-1 at adjacent position Ala66, which is known to participate as a half-cystine with position 83 in FGF-8, FGF-19, and FGF-23. Results show that, unlike position 83, a free cysteine at position 66 destabilizes FGF-1; however, upon oxidation, a near-optimal disulfide bond is formed between Cys66 and Cys83, resulting in approximately 14 kJ/mol of increased thermostability. Thus, while the conserved free cysteine at position 83 in the majority of the FGF proteins may have a principal role in limiting functional half-life, evidence suggests that it is a vestigial half-cystine.
Lee
Jihun
J
Department of Biomedical Sciences, College of Medicine, Florida State University, Tallahassee, FL 32306-4300, USA.
Blaber
Michael
M
D019943:Amino Acid Substitution; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000235:genetics*; D000089703:Fibroblast Growth Factor-23; D005346:Fibroblast Growth Factors / Q000737:chemistry* / Q000235:genetics*; D006801:Humans; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011489:Protein Denaturation; D055550:Protein Stability; D017434:Protein Structure, Tertiary
C000717427:FGF23 protein, human; C000717428:Fgf23 protein, mouse; D005346:Fibroblast Growth Factors; D000089703:Fibroblast Growth Factor-23; D003545:Cysteine