The Recombinant Expression, Purification And Activity Analysis Of LDH Of Veillonella Dispar

Veillonella is a kind of Gram-negative coccus, it mainly parasitize in oral, intestinal canal and respiratory tract of animals and human. Veillonella has ability of acid resistance to some extant and it can grow in acid circumstances.Veillonella plays a very delicate role in dental plaque. It can change the intense acid lactate to the weak acid but when it is catalyzed by sugar it can also lead to the huge production of lactate . Veillonella has this function all because of lactate dehydrogenase(LDH) . LDH is a kind of constitutive enzyme of bacteria. It can be put into different categories as dependence-LDH ( nLDH ) and independence-LDH (iLDH) which depend on it whether depend Nicotinamide adenine dinucleotide phosphate (NAD).nLDH plays this role. The concrete function and mechanism of action of this gene is not so clear presently. But it is affirmative that Veillonella is interpersonally connected to the happen of dental caries and the grow of dental plaque. So it is very necessary to carry on deep research in Veillonella.In earlier period of experiment LDH in Veillonella had been successfully cloned and preliminary expression .At the same time the recombinant plasmid pET-28a-LDH had been constructed. The sequence of LDH in Veillonella has login American GenBank and the sequence number is EU518464.This experiment transform the recombinant plasmid pET-28a-LDH in BL21(DE3) and Rosetta(DE3) and efficiently express the protein respectively. The expression of recombinant plasmid pET-28a-LDH in BL21(DE3) exceed which in Rosetta(DE3). We also carry on the character of goal protein to appraise. The SDS-PAGE electrophoresis display that the obvious protein expression band at 33KD. After the purification by HisTRAP FF column LDH reactive kit determine that the recombinant protein has activity. This experiment lay the foundation of continued study of LDH in Veillonella from the biological perspective and research the new path of dental caries prevention.