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Eukaryotic in vitro translation systems require large numbers of protein and RNA components and thereby rely on the use of cell extracts. Here we established a new in vitro translation system based on rice callus extract (RCE). We confirmed that RCE maintains its initial activity even after five freeze-thaw cycles and that the optimum temperature for translation is around 20°C. We demonstrated that the RCE system allows the synthesis of hERG, a large membrane protein, in the presence of liposomes. We also showed that the introduction of a bicistronic mRNA based on 2A peptide to RCE allowed the production of two distinct proteins from a single mRNA. Our new method thus facilitates laboratory-scale production of cell extracts, making it a useful tool for the in vitro synthesis of proteins for biochemical studies. A novel method for preparing extracts of rice cultured cells for the establishment of a cell-free protein expression system [ABSTRACT FROM AUTHOR]

Glucolipids inBacillus subtilisare synthesized by UgtP processively transferring glucose from UDP-glucose to diacylglycerol. Here we conclude that the abnormal morphology of augtPmutant is caused by lack of glucolipids, since the same morphology arises after abolition of glucolipid production by disruption ofpgcAandgtaB,which are involved in UDP-glucose synthesis. Conversely, expression of a monoglucosyldiacylglycerol (MGlcDG) produced by 1,2-diacylglycerol 3-glucosyltransferase fromAcholeplasma laidlawii(alMGS) almost completely suppressed theugtPdisruptant phenotype. Activation of extracytoplasmic function (ECF) sigmas (SigM, SigV, and SigX) in theugtPmutant was decreased by alMGS expression, and was suppressed to low levels by MgSO4addition. When alMGS and alDGS (A. laidlawii1,2-diacylglycerol-3-glucose (1-2)-glucosyltransferase producing diglucosyldiacylglycerol (DGlcDG)) were simultaneously expressed, SigX activation was repressed to wild type level. These observations suggest that MGlcDG molecules are required for maintenance ofB. subtiliscell shape and regulation of ECF sigmas, and DGlcDG regulates SigX activity. The abnormal morphology of theB. subtilis ugtPmutant was suppressed by expression of heterologous MGlcDG synthase (left). Effect of heterologous glucolipids on activitiy of ECF sigmas (right). [ABSTRACT FROM AUTHOR]

The article focuses on a study that characterizes the nuclear-and plastid-encoded SecA-homologous genes in red alga Cyanidioschyzon merolae. It states that SecA is an adenosine triphosphate (ATP)-driven motor for protein translocation found in plants and bacteria wherein listeria and mycobacteria were found to possess such. It adds that the red alga has also possessed the genes, one encoded in the cell nucleus and one in the plastid genome, playing varied roles in protein translocation.

The article examines the heterologous expression of the Oceanobacillus iheyensis sigma factor SigW and its anti-protein RsiW in Bacillus subtilis. It explores the protein structure and gene organization of extra-cytoplasmic-function (EFC) sigma factors in the cytoplasmic membrane and periplasmic space and their genetic competence in manipulating the genome of Bacillus subtilis. It also notes the need to consider the mechanisms on protein translocation in designing heterologous gene expression.

The article presents a study which examines the nucleotide sequence of the genome of Bacillus subtilis pseudolysogenic phage SP10 and the comparison between SP10 with virulent phage SPO1 based on their gene organization and regulation. It explores the resemblances and differences of the SP10 and SPO1 genomes as to their sensitivity to restriction enzymes in base modification. It also looks into the enzyme activity of the genomes in host genes and the phage genes.