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pH-dependent association of factor VIII chains: enhancement of affinity at physiological pH by Cu2+.

Authors :
Wakabayashi H
Zhou Q
Nogami K
Ansong C
Varfaj F
Miles S
Fay PJ
Source :
Biochimica et biophysica acta [Biochim Biophys Acta] 2006 Jun; Vol. 1764 (6), pp. 1094-101. Date of Electronic Publication: 2006 Apr 22.
Publication Year :
2006

Abstract

Reconstitution of factor VIII from isolated heavy chain (HC) and light chain (LC) shows pH-dependence. In the presence of Ca2+, up to 80% of native factor VIII activity was recovered over a wide range of pH. In contrast, affinity of HC and LC was maximal at pH 6.5-6.75 (Kd approximately 4 nM), whereas a Kd approximately 20 nM was observed at physiological pH (7.25). The effect of Cu2+ (0.5 microM total Cu2+) on maximal activity regenerated was negligible at pH 6.25-8.0. However, this level of Cu2+ increased the inter-chain affinity by approximately 5-fold at pH 7.25. This effect resulted from an approximately 1.5-fold increased association rate constant (k(on)) and an approximately 3-fold reduced dissociation rate constant (k(off)). High affinity (Kd=5.3 fM) of the factor VIII heterodimer for Cu2+ was estimated by increases in cofactor activity. No significant increase in inter-chain affinity was observed when either isolated chain was reacted with Cu2+ followed by addition of the complementary chain. Together, these results suggest that the protonation state of specific residues modulates inter-chain affinity. Furthermore, copper ion contributes to the maintenance of the heterodimer at physiologic pH by a mechanism consistent with bridging the two chains.

Details

Language :
English
ISSN :
0006-3002
Volume :
1764
Issue :
6
Database :
MEDLINE
Journal :
Biochimica et biophysica acta
Publication Type :
Academic Journal
Accession number :
16731058
Full Text :
https://doi.org/10.1016/j.bbapap.2006.04.004