Crystallization of antiangiogenic Kringle V derived from human apolipoprotein A: crystallization applied to purification and formulation.
Biosci Biotechnol Biochem
; 70(4): 916-25, 2006 Apr.
Article
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| MEDLINE
| ID: mdl-16636459
ABSTRACT
In this study, the Kringle V domain (Glu4225-Ser4310) of human apolipoprotein A, an antiangiogenic polypeptide, was expressed as a secreted form in Pichia pastoris, and was purified via a process consisting of three chromatographic steps. The chromatographically purified kringle V domain contained a C-terminal serine-deleted form and several high-molecular-weight forms, which were suspected to represent glycosylated derivatives. In order to remove these derivatives, we employed a crystallization process. The crystallization of kringle V resulted in an 85% recovery yield, and also resulted in the complete removal of the aforementioned high-molecular-weight forms. However, we were still able to detect a trace of the C-terminal serine-deleted form. The prepared Kringle V crystals were stable within a pH range of 7.0 to 8.0, and were completely dissolved by dilution, which is a crucial factor in the preparation of a highly concentrated formulation. The chromatogram of the crystallized kringle V on reversed-phase HPLC analysis was identical to that observed without crystallization. Also, we noted that the original anti-wound migration activities of the molecule toward human umbilical vein endothelial cells were completely retained.
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Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Apolipoproteínas A
/
Inhibidores de la Angiogénesis
Límite:
Humans
Idioma:
En
Revista:
Biosci Biotechnol Biochem
Asunto de la revista:
BIOQUIMICA
/
BIOTECNOLOGIA
Año:
2006
Tipo del documento:
Article