The clear presence of serine protease inhibitors has been detected in organisms and in animal and plant cells. This study describes the isolation and characterization of a Kunitz form inhibitor from G. dubium seed extract, which showed action against bovine trypsin and chymotrypsin. Raf inhibition This is the first trypsin inhibitor which also has lectin like properties. Initially, affinity chromatography on a thyroglobulinagarose line was useful for purification, with the intention of obtaining a lectin. When the isolated protein was recognized as a trypsin inhibitor, an alternative solution purification process, involving affinity chromatography on a trypsinagarose line, allowed the preparation of the same material with a far greater yield. With both processes, the fraction obtained showed exactly the same two bands in SDS PAGE, of 20,000 and 22,000 apparent molecular weights, which could not be resolved by reverse phase HPLC or by Mono Q or MonoS Lonafarnib ic50 chromatography and which showed only one band on native PAGE. The amino terminal sequence of these companies was similar. Furthermore, by trypsin digestion followed by mass spectrometry, 16 proteins were found to own identical mass. All these findings strongly declare that they are closely related proteins. The different mobility on SDSPAGE could possibly be due to posttranslational modifications near the C terminus or even to a glycosylation pattern, though in these cases they’d have now been expected to split by some of the chromatographic techniques assayed. To clarify this point, PAS staining of SDSPAGE was conducted, confirming that the 22 kDa band is glycosylated. Furthermore, Cholangiocarcinoma molecular mass of PDTI was based on MALDI TOF MS, showing two main peaks of approximately 18 and 20 kDa. Size exclusion chromatography unveiled that PDTI behaves as a monomeric protein. This research was carried out both in the presence and in the absence of Ca2t, to avoid the possible interaction of PDTI with the column matrix, which may lead to underestimation of its native molecular mass, taking into consideration that carbohydrate binding of PDTI is Ca2t dependent. Because of the high level of amino terminal sequence identification of PDTI with Kunitz type trypsin inhibitors, trypsin and chymotrypsin inhibitory activities of PDTI were calculated and the particular Ki values determined. It was found to really have a greater appreciation for trypsin than for chymotrypsin. The lectin like qualities of PDTI were proved by its hemagglutinating activity on trypsin addressed rabbit erythrocytes, in the presence of Ca2t. When SBTI was tried in exactly the same assay, it was found to generally share this hemagglutinating activity. Even though SBTI has been carefully studied, this property had remained undetected, order Docetaxel probably due to its failure to agglutinate human erythrocytes and to the need of Ca2t in the medium.