Waffenschmidt S, Kusch T, Woessner J P
Institut für Biochemie, Universität zu Köln, Köln, Germany.
Plant Physiol. 1999 Nov;121(3):1003-15. doi: 10.1104/pp.121.3.1003.
The addition of primary amines to the growth medium of the unicellular green alga Chlamydomonas reinhardtii disrupts cell wall assembly in both vegetative and zygotic cells. Primary amines are competitive inhibitors of the protein-cross-linking activity of transglutaminases. Two independent assays for transglutaminase confirmed a burst of extracellular activity during the early stages of cell wall formation in both vegetative cells and zygotes. When non-inhibiting levels of a radioactive primary amine ((14)C-putrescine) were added to the growth medium, both cell types were labeled in a reaction catalyzed by extracellular transglutaminase. The radioactive label was found specifically in the cell wall proteins of both cell types, and acid hydrolysis of the labeled material released unmodified (14)C-putrescine. Western blots of the proteins secreted at the times of maximal transglutaminase activity in both cell types revealed a single highly cross-reactive 72-kD band when screened with antibodies to guinea pig tissue transglutaminase. Furthermore, the proteins immunoprecipitated by this antiserum in vivo exhibited transglutaminase activity. We propose that this transglutaminase is responsible for an early cell wall protein cross-linking event that temporally precedes the oxidative cross-linking mediated by extracellular peroxidases.
向单细胞绿藻莱茵衣藻的生长培养基中添加伯胺会破坏营养细胞和合子细胞中的细胞壁组装。伯胺是转谷氨酰胺酶蛋白质交联活性的竞争性抑制剂。两种独立的转谷氨酰胺酶检测方法证实，在营养细胞和合子的细胞壁形成早期阶段，细胞外活性会出现爆发。当向生长培养基中添加非抑制水平的放射性伯胺（（14）C-腐胺）时，两种细胞类型都会在细胞外转谷氨酰胺酶催化的反应中被标记。在两种细胞类型的细胞壁蛋白中都特异性地发现了放射性标记，并且对标记物质进行酸水解后释放出未修饰的（14）C-腐胺。用针对豚鼠组织转谷氨酰胺酶的抗体进行筛选时，在两种细胞类型中转谷氨酰胺酶活性最高时分泌的蛋白质的蛋白质免疫印迹显示出一条单一的高度交叉反应性的72-kD条带。此外，该抗血清在体内免疫沉淀的蛋白质表现出转谷氨酰胺酶活性。我们认为这种转谷氨酰胺酶负责在时间上先于细胞外过氧化物酶介导的氧化交联的早期细胞壁蛋白交联事件。
