Progresses in predicting the crystallizability of proteins / 生物工程学报

Determination of protein 3-dimensional structure offers very important information in biology researches, especially for understanding protein functions and redundant drug design. The X-ray crystallography is still the main technique for protein structure determination. Obtaining protein crystals is an essential procedure after protein purification in this technique. However, there is only 42% of soluble purified proteins yield crystals by statistics. Experimental verification of protein crystallizability is relatively expensive and time-consuming. Thus it is desired to predict the protein crystallizability by a computational method before starting the experiment. In this paper, combined with our own efforts, some successful in silico methods to predict the protein crystallizability are reviewed.

Expression of human aspartyl beta-hydroxylase and preparation of its monoclonal antibody / 生物工程学报

We investigated the mechanism of human aspartyl beta-hydroxylase (HAAH) in early diagnosis of tumors. The encoding gene of HAAH was cloned from the hepatic carcinoma by RT-PCR and expressed as a fused protein in the prokaryotic vector pBV-IL1. The expressed HAAH was purified by Ni(2+)-NTA purification column and the purified protein was then used to immunize Balb/c mice. Three hybridoma cell lines (respectively designated H3/E10, E4/F12 and G4/D8) stably expressing the monoclonal antibody specific to HAAH fusion protein were obtained. The specificity and sensitivity of the monoclonal antibody were assessed by indirect enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. Finally, the monoclonal antibody expressed by H3/E10 cell line was used to detect the expression of HAAH in several tumor cell lines by indirect immuno-fluorescence, and the specific fluorescence was observed. In conclusion, this study successfully constructed the recombinant prokaryotic vector pBV-IL1-HAAH and prepared HAAH-specific monoclonal antibody for further study of the structure and function of the protein. The result may also lay solid foundation for the research of the molecular mechanism of HAAH in early diagnosis of tumors.