Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.6 (thromboplastin)
 13,278 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Decorin is a small leucine-rich proteoglycan which is a component of the extracellular matrix of many connective tissues. We have developed a method for expression and purification of decorin as a fusion protein in Escherichia coli. A PCR product coding for the 330-amino-acid-residue secreted form of bovine decorin core protein was cloned into the vector pMal-c for expression in E. coli as a maltose-binding protein (MBP) fusion. Expressed MBP-decorin tended to form insoluble aggregates resistant to degradation by E. coli intracellular proteases. Fusion protein was therefore solubilized from bacterial inclusion bodies using guanidine HCl and refolded by dilution of the chaotrope to minimally denaturing conditions and disulfide shuffling. Final purification included amylose resin affinity chromatography and size exclusion chromatography. The 79-kDa MBP-decorin fusion protein could be completely cleaved by factor Xa protease in 24 h to yield 43-kDa MBP and 36-kDa decorin core protein. The decorin core protein was separated from MBP and factor Xa protease by DEAE-Sephacel chromatography. Using a solid-phase assay, we have characterized its binding affinity for extracellular matrix components known to interact with tissue-derived decorin including collagen type VI, collagen type I, and fibronectin. The purification and refolding protocol described here may be generally applicable to bacterial expression of other members of this growing family of related small leucine-rich proteoglycan core proteins.
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PMID:Purification and characterization of decorin core protein expressed in Escherichia coli as a maltose-binding protein fusion. 881 84

Secreted phospholipases A(2) (sPLA(2)s) form a large family of structurally related enzymes which are widespread in nature. Snake venoms are known for decades to contain a tremendous molecular diversity of sPLA(2)s which can exert a myriad of toxic and pharmacological effects. Recent studies indicate that mammalian cells also express a variety of sPLA(2)s with ten distinct members identified so far, in addition to the various other intracellular PLA(2)s. Furthermore, scanning of nucleic acid databases fueled by the different genome projects indicates that several sPLA(2)s are also present in invertebrate animals like Drosophila melanogaster as well as in plants. All of these sPLA(2)s catalyze the hydrolysis of glycerophospholipids at the sn-2 position to release free fatty acids and lysophospholipids, and thus could be important for the biosynthesis of biologically active lipid mediators. However, the recent identification of a variety of membrane and soluble proteins that bind to sPLA(2)s suggests that the sPLA(2) enzymes could also function as high affinity ligands. So far, most of the binding data have been accumulated with venom sPLA(2)s and group IB and IIA mammalian sPLA(2)s. Collectively, venom sPLA(2)s have been shown to bind to membrane and soluble mammalian proteins of the C-type lectin superfamily (M-type sPLA(2) receptor and lung surfactant proteins), to pentraxin and reticulocalbin proteins, to factor Xa and to N-type receptors. Venom sPLA(2)s also associate with three distinct types of sPLA(2) inhibitors purified from snake serum that belong to the C-type lectin superfamily, to the three-finger protein superfamily and to proteins containing leucine-rich repeats. On the other hand, mammalian group IB and IIA sPLA(2)s can bind to the M-type receptor, and group IIA sPLA(2)s can associate with lung surfactant proteins, factor Xa and proteoglycans including glypican and decorin, a mammalian protein containing a leucine-rich repeat.
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PMID:Increasing molecular diversity of secreted phospholipases A(2) and their receptors and binding proteins. 1108 Jun 77