title: Lipid binding region (2303-2332) is involved in aggregation of recombinant human FVIII (rFVIII).
abstract: Factor VIII (FVIII) is a multi-domain protein that is important in the clotting cascade. Its deficiency causes Hemophilia A, a bleeding disorder. The unfolding of protein domains can lead to physical instability such as aggregation, and hinder their use in replacement therapy. It has been shown that the aggregation of rFVIIII is initiated by small fluctuations in the protein's tertiary structure (Grillo et al., 2001, Biochemistry 40:586-595). We have investigated the domain(s) involved in the initiation of aggregation using circular dichroism (CD), size exclusion chromatography (SEC), fluorescence anisotropy, domain specific antibody binding, and clotting activity studies. The studies indicated that aggregation may be initiated as a result of conformational change in the C2 domain encompassing the lipid-binding region (2303-2332). The presence of O-phospho-L-Serine (OPLS), which binds to the lipid-binding region of FVIII, prevented aggregation of the protein.
authors.affiliation: Department of Pharmaceutical Sciences, University at Buffalo, State University of New York, Amherst, NY 14260, USA.
authors.forename: Karthik
authors.initials: K
authors.lastname: Ramani
authors.forename: Vivek S
authors.initials: VS
authors.lastname: Purohit
authors.forename: Razvan D
authors.initials: RD
authors.lastname: Miclea
authors.forename: C Russell
authors.initials: CR
authors.lastname: Middaugh
authors.forename: Sathyamangalam V
authors.initials: SV
authors.lastname: Balasubramanian
mesh_terms: D001665:Binding Sites; D005169:Factor VIII / Q000737:chemistry*; D050356:Lipid Metabolism; D010446:Peptide Fragments / Q000737:chemistry*; D011487:Protein Conformation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry; D013696:Temperature
chemical_list: D010446:Peptide Fragments; D011994:Recombinant Proteins; D005169:Factor VIII