Cloning and expression of cytoskeleton-related gene mreB and expression differences between two shapes of Spiroplasma melliferum

Abstract:

[Objective] mreB1-5, the cytoskeleton-related genes of Spiroplasma melliferum CH-1 were cloned and expressed in Escherichia coli. Then their proteins properties were predicted. Expression levels of these genes in spiral and non-spiral shape of spiroplasma were determined. [Methods] Coding sequences of mreB1-5 were amplified from the genomic DNA of S. melliferum CH-1. Then recombination plasmids pETmreB1-5 were expressed in E. coli BL21 respectively after IPTG induction. Ni-NTA was used to purify the recombinant proteins. The properties of MreB proteins were predicted by online tools. Real-Time quantitative PCR method was applied to compare the expression levels of mreB1-5 genes between two obvious different shapes of S. melliferum CH-1. [Results] Five mreB genes from S. melliferum were cloned. The molecular weight of five MreB proteins were 36, 23, 23, 37, 25 kD, respectively. They were possibly hydrophobic proteins with no signal peptide and were identified as part of MreB/Mbl protein family. The results exhibited that the expression levels of genes mreB1-5 in non-spiral shape of spiroplasma were relatively lower than that in spiral shape. [Conclusion] This is the first report of clone and expression of spiroplasma cytoskeleton-related genes mreB1-5 which encode actin-like protein MreB. It could be inferred that these genes are closely related with spiroplasma morphology. All results contribute to further investigate the function of actin-like protein MreB in spiral shape and movement of spiroplasma.