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Lectins are (glyco) proteins of non-immune origin, which specifically bind carbohydrates without modifying them. Although lectins have been found in many organisms, plant lectins are probably the most well known. This unique group of proteins has provided researchers with powerful tools to explore a myriad of biological structures and processes. In the current work, the DNA fragment coding for the mature protein of the seed lectin from Pterocarpus indicus and Pterocarpus echinatus (family: Leguminosae, subfamily: Papilionoideae, tribe: Dalbergieae) was cloned into the expression vector pBADMycHis and transformed into E.coli TOP10 cells. Expression in TOP10 cells and purification of the recombinant lectin from P. echinatus was successfully achieved. Nevertheless, expression and purification of the recombinant P. indicus lectin only yielded extremely low amounts of purified protein. From our experiments it is estimated that around 1 and 7.6 mg of recombinant P. indicus and P. echinatus lectin respectively could be purified from 1 liter of bacterial culture after induction with arabinose. The expressed protein has the same molecular weight as that of the seed lectin determined previously and shows also similar but not identical carbohydrate-binding properties. As for the seed lectin, both recombinant lectins are specific for mannose and glucose and do not bind galactose. In the case of the recombinant P. indicus lectin, mannose seems to be a stronger inhibitor when comparing the seed lectin. Unlike seed lectin, recombinant P. indicus lectin has affinity for mucin type I and peroxidase, as well as less affinity for fetuin and higher affinity for thyroglobulin and trypsin inhibitor. In the case of the recombinant P. echinatus lectin, the only remarkable difference is that methyl-alpha,D-glucopyranoside is a more potent inhibitor of agglutination than for the seed lectin, whereas ovalbumin seems to better inhibit the seed lectin than the recombinant protein. The differences in affinity of both recombinant and seed lectins can be explained as the recombinant lectin is probably one of the isolectins purified from the seed. It is also possible that the difference is due to protein glycosylation. The expression and purification of lectins in heterologous systems has the advantage of the availability of lectin molecules independent of the availability of plant material. Moreover, it allows studies on mutagenesis and the purification of only one isolectin from the mixture of isolectins that is extracted from the seeds. |
