The Cystine Knot Promotes Folding and Not Thermodynamic Stability in Vascular Endothelial Growth Factor
Cystine knots consist of three intertwined disulfide bridges and are considered major determinants of protein stability in proteins in which they occur. We questioned this function and observed that removal of individual disulfide bridges in human vascular endothelial growth factor (VEGF) does not r...
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Published in | The Journal of biological chemistry Vol. 277; no. 45; pp. 43410 - 43416 |
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Main Authors | , , , , |
Format | Journal Article |
Language | English |
Published |
United States
American Society for Biochemistry and Molecular Biology
08.11.2002
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Subjects | |
Online Access | Get full text |
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Summary: | Cystine knots consist of three intertwined disulfide bridges and are considered major determinants of protein stability in
proteins in which they occur. We questioned this function and observed that removal of individual disulfide bridges in human
vascular endothelial growth factor (VEGF) does not reduce its thermodynamic stability but reduces its unexpected high thermal
stability of 108â°C by up to 40â°C. In wild-type VEGF ( ÎG
= 5.1 kcal·mol â1 ), the knot is responsible for a large entropic stabilization of T Î S
= â39.3 kcal mol â1 , which is compensated for by a ÎH
of â34.2 kcal mol â1 . In the disulfide-deficient mutants, this entropic stabilization disappears, but instead of a decrease, we observe an increase
in the thermodynamic stability by about 2 kcal·mol â1 . A detailed crystallographic analysis of the mutant structures suggests a role of the cystine knot motif in protein folding
rather than in the stabilization of the folded state. When assuming that the sequential order of the disulfide bridge formation
is conserved between VEGF and glycoprotein α-subunit, the crystal structure of the mutant C61A-C104A, which deviates by a
root mean square deviation of more than 2.2 Ã
from wild-type VEGF , identifies a true folding intermediate of VEGF . |
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ISSN: | 0021-9258 1083-351X |
DOI: | 10.1074/jbc.M206438200 |