ABSTRACT
Co-operation between SMEs and Academia can be a win-win situation when each partner understands the constraints of the other. SMEs are often leaders in innovation; therefore more ready to share interest in research. They are flexible and dynamic. They need a short feed-back to sustain their co-operation. Academia is often more long-term oriented and more question- than answer-oriented. A code of conduct can ease the relationship because it can anticipate the potential problems.
Subject(s)
Academies and Institutes , Commerce , Infant Food/standards , Cooperative Behavior , Focus Groups , Humans , Infant , Infant, NewbornABSTRACT
BACKGROUND: Carboxypeptidase U (EC 3.4.17.20, TAFIa) is a new member of the metallocarboxypeptidase family circulating in human plasma as a zymogen. It is activated during coagulation and is considered as an important player in the regulation of fibrinolysis. METHODS: Heterologous expression of human plasma procarboxypeptidase U (proCPU, TAFI) was obtained in mammalian cells (C127 and DON) and in insect cells (Sf21 and H5 cells). Conditioned media were purified by cation-exchange chromatography and plasminogen affinity chromatography to yield an essentially pure protein. RESULTS: All systems gave high expression levels (6-20 mg/l). Due to differences in glycosylation of the activation peptide, the recombinant variants of proCPU migrated differently on SDS-PAGE (52-65 kDa). However, after activation, all active recombinant enzymes migrated at 35 kDa, similar to native CPU and no evidence for post-translational modification of the catalytic domains could be detected. For the mammalian cell produced variants, activation was more efficient after desialylation. After activation, CPU showed low solubility (0.2 mg/ml) but was inhibited similarly as native CPU. CONCLUSIONS: Mammalian cell systems were the most efficient for the production of human plasma recombinant proCPU. The obtained zymogen differs with respect to the extent and the heterogeneity of glycosylation but, after activation, the experiments did not reveal any alteration between the recombinant and native protein.
