Department of Biological Sciences, Oakwood University, Huntsville, AL 35896, USA.
Department of Basic Sciences, Division of Microbiology and Molecular Genetics, School of Medicine, Loma Linda University, Loma Linda, CA 92350, USA.
Microbiology (Reading). 2010 Oct;156(Pt 10):3065-3072. doi: 10.1099/mic.0.038315-0. Epub 2010 Jul 1.
Recombinant VimA protein can interact with the gingipains and several other proteins that may play a role in its biogenesis in Porphyromonas gingivalis. In silico analysis of PG2096, a hypothetical protein that was shown to interact with VimA, suggests that it may have environmental stress resistance properties. To further evaluate the role(s) of PG2096, the predicted open reading frame was PCR amplified from P. gingivalis W83 and insertionally inactivated using the ermF-ermAM antibiotic-resistance cassette. One randomly chosen PG2096-defective mutant created by allelic exchange and designated FLL205 was further characterized. Under normal growth conditions at 37 °C, Arg-X and Lys-X gingipain activities in FLL205 were reduced by approximately 35 % and 21 %, respectively, compared to the wild-type strain. However, during prolonged growth at an elevated temperature of 42 °C, Arg-X activity was increased by more than 40 % in FLL205 in comparison to the wild-type strain. In addition, the PG2096-defective mutant was more resistant to oxidative stress when treated with 0.25 mM hydrogen peroxide. Taken together these results suggest that the PG2096 gene, designated regT (regulator of gingipain activity at elevated temperatures), may be involved in regulating gingipain activity at elevated temperatures and be important in oxidative stress resistance in P. gingivalis.
重组 VimA 蛋白可以与牙龈卟啉单胞菌中的牙龈蛋白酶和其他几种可能参与其生物发生的蛋白质相互作用。对与 VimA 相互作用的假定蛋白 PG2096 进行计算机分析表明，它可能具有环境应激抗性。为了进一步评估 PG2096 的作用，从牙龈卟啉单胞菌 W83 中扩增预测的开放阅读框，并使用 ermF-ermAM 抗生素抗性盒插入失活。通过等位基因交换随机创建的一个 PG2096 缺陷突变体 FLL205，并进一步进行了表征。在 37 °C 的正常生长条件下，与野生型菌株相比，FLL205 中的 Arg-X 和 Lys-X 牙龈蛋白酶活性分别降低了约 35%和 21%。然而，在 42 °C 的高温下长时间生长时，与野生型菌株相比，FLL205 中的 Arg-X 活性增加了 40%以上。此外，与野生型菌株相比，PG2096 缺陷突变体在用 0.25 mM 过氧化氢处理时对氧化应激更具抗性。总之，这些结果表明，PG2096 基因（命名为 regT，即高温下牙龈蛋白酶活性的调节剂）可能参与调节高温下的牙龈蛋白酶活性，并在牙龈卟啉单胞菌的氧化应激抗性中起重要作用。
