ObjectiveTo evaluate the expression of endothelial differentiation gene/lysophosphatidic acid (LPA) receptors (Edg/LPA) and its clinical significance in human pancreatic cancer.MethodsFifty cases of pancreatic cancer and adjacent normal tissues were collected,and Real-time PCR,Western blot and immunohistochemistry was used to determine the expression of Edg-2/LPA1,Edg-4/LPA2 and Edg-7/LPA3 receptors mRNA and protein,and its relationship with clinicopathological parameters was analyzed.Results The expressions of Edg-2/LPA1,Edg-4/LPA2,Edg-7/LPA3 receptor mRNA were (0.142 ± 0.042 ) ％,(0.471 ±0.064)％,(0.231 ±0.043)％ in pancreatic cancer,and the corresponding values were (0.132 ±0.029)％,(0.027 ±0.015)％,(0.163 ±0.046)％ in adjacent normal tissues.The expressions of Edg-4/LPA2 receptor mRNA in pancreatic cancer were significantly lower than that in adjacent normal tissues ( P ＜0.05 ).The expressions of Edg-4/LPA2 receptor protein in pancreatic cancer were significantly lower than that in adjacent normal tissues ( P ＜ 0.05 ).The expressions of three types of Edg /LPA receptor mRNA in pancreatic cancer were parallel to serum CA19-9 levels.The expressions of Edg-4/LPA2 receptor mRNA were associated with tumor size,differentiation degree,and invasive ability and metastasis.While the expressions of Edg-2/LPA1,Edg-7/LPA3 receptor mRNA was associated with invasive ability and metastasis only.ConclusionsEdg-4/LPA2 receptor is highly expressed in pancreatic cancer,which suggesting the malignant biological behavior of pancreatic cancer.

To investigate the mechanism of high product specificity of gamma-clodextrin glucanotransferase (CGTase) from alkalophilic Bacillus clarkii 7364, we aligned protein sequence and structure model, found out that loss of 6 amino acids at subsite -7 probably affected its product specificity. Using overlapping PCR method, we inserted 6 amino acids into subsite -7 of CGTase. The mutant CGTase gene was ligated with pET-20b (+) and expressed in Escherichia coli BL21 (DE3). The extracellular recombinant enzyme was used to transform soluble starch into cyclodextrins (CDs). HPLC analysis results show that, compared to wild CGTase, the gamma-CDs produced by mutant enzyme decreased from 76.0% to 12.5%, whereas the ratio of alpha- and beta-CDs increased from 8.7% and 15.2% to 37.5% and 50%. The possible mechanism was that, compared to alpha-, beta-CGTase, wild gamma-CGTase lacks 6 amino acids in its subsite -7. This conformation provided more space for glucose combination and was thus advantageous for forming gamma-CD. When the 6 amino acids were inserted into the subsite -7 of wild gamma-CGTase, the space to bind with glucose reduced and consequently resulted in less gamma-CD production.
Bacillus ; enzymology ; genetics ; Escherichia coli ; genetics ; metabolism ; Glucosyltransferases ; biosynthesis ; genetics ; Mutant Proteins ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics