Biology and Chemistry Department, Benedict College, Columbia, SC 29204, USA.
Protein J. 2013 Aug;32(6):426-34. doi: 10.1007/s10930-013-9499-9.
Tissue transglutaminase (TGC, TG2, 80 kDa) is inactive in cross-linking reactions and is converted in vitro and in vivo to the TG (55 kDa) active isoform (Fraij in J Cell Biochem 112:2469-2489, 2011). Two isoforms of human TGC were cloned from human erythroleukemia (HEL) cells induced with retinoic acid (RA) and termed TGH, 63 kDa (Fraij et al. in J Biol Chem 267:22616-22673, 1992) and TGH2, 37 kDa. The purified TGC isoforms exhibited GTPase activity and TGH and TGH2 showed higher activities than the native TGC protein. In all normal cells examined, TGC was found in membrane fractions several fold higher than the supernatant fractions; however, in the natural tumor cell line HEL the TGC cellular distribution was reversed. Although TGC is the major enzyme in normal human erythrocytes, its expression level was significantly decreased in HEL cells. RA treatment induced a sevenfold increase in the level of TGC protein in HEL cells and was accompanied by its translocation to cell membranes. When isolated membrane and supernatant fractions from normal human foreskin (CF3), normal human embryonic lung (WI-38), and HEL cells treated with or without RA were incubated with [(32)P]-ATP at 37 °C for 1 h, more radio-labeled proteins were detected in the membrane fractions than the cytosolic fractions. More labeled protein bands were detected in RA treated HEL cells in comparison to control HEL cell extracts. Radio labeled proteins coimmunoprecipitated with the TGC isoforms in RA treated HEL membrane fractions thereby confirming that the radio-labeled material consists of endogenous proteins associated with TGC isoforms. Protein phosphorylation is related to the induction and translocation of the isoforms in RA treated cells. These results show that the TGC isoforms complexes with proteins in vivo and that the phosphorylation of these proteins is catalyzed directly by TGC kinase activity or indirectly by the TGC phosphorylation of other protein kinases.
组织转谷氨酰胺酶（TGC、TG2、80 kDa）在交联反应中无活性，在体外和体内转化为 TG（55 kDa）活性同工型（Fraij 在 J Cell Biochem 112：2469-2489，2011）。从维甲酸（RA）诱导的人红白血病（HEL）细胞中克隆了两种人类 TGC 同工型，分别命名为 TGH，63 kDa（Fraij 等人在 J Biol Chem 267：22616-22673，1992）和 TGH2，37 kDa。纯化的 TGC 同工型表现出 GTPase 活性，TGH 和 TGH2 的活性高于天然 TGC 蛋白。在所有检查的正常细胞中，TGC 在膜部分的含量比上清部分高几倍；然而，在天然肿瘤细胞系 HEL 中，TGC 的细胞分布是相反的。尽管 TGC 是正常人类红细胞中的主要酶，但在 HEL 细胞中的表达水平显著降低。RA 处理诱导 HEL 细胞中 TGC 蛋白水平增加了七倍，并伴随着其向细胞膜的易位。当从正常人类包皮（CF3）、正常人类胚胎肺（WI-38）和用或不用 RA 处理的 HEL 细胞中分离的膜和上清部分在 37°C 下用 [(32)P]-ATP 孵育 1 小时时，在膜部分中检测到比胞质部分更多的放射性标记蛋白。与对照 HEL 细胞提取物相比，在 RA 处理的 HEL 细胞中检测到更多标记的蛋白带。在用 RA 处理的 HEL 膜部分中，放射性标记的蛋白质与 TGC 同工型共免疫沉淀，从而证实放射性标记的物质由与 TGC 同工型相关的内源性蛋白质组成。蛋白质磷酸化与 RA 处理细胞中同工型的诱导和易位有关。这些结果表明 TGC 同工型在体内与蛋白质形成复合物，并且这些蛋白质的磷酸化是由 TGC 激酶活性直接催化的，或者是由 TGC 对其他蛋白激酶的磷酸化间接催化的。
