HU is a most abundant DNA-binding protein in bacteria. This protein is conserved either in its heterodimeric form or in one of its homodimeric forms in all bacteria, in plant chloroplasts, and in some viruses. HU protein non-specifically binds and bends DNA as a hetero- or homodimer and can participate in DNA supercoiling and DNA condensation. It also takes part in some DNA functions such as replication, recombination, and repair. HU does not recognize any specific sequences but shows some specificity to cruciform DNA and to repair intermediates, e.g., nick, gap, bulge, 3'-overhang, etc. To understand the features of HU binding to DNA and repair intermediates, a fast and easy HU proteins purification procedure is required. Here we report overproduction and purification of the HU homodimers. The method of HU purification allows obtaining a pure recombinant non-tagged protein cloned in Escherichia coli. We applied this method for purification of Acholeplasma laidlawii HU and demonstrated that this protein possesses a DNA-binding activity and is free of contaminating nuclease activity. Besides that we have shown that expression of A. laidlawii ihf_hu gene in a slow-growing hupAB E. coli strain restores the wild-type growth indicating that aclHU can perform the basic functions of E. coli HU in vivo.
Matched MeSH terms: Escherichia coli/growth & development
The spontaneous occurrence of resistance to the herbicide glyphosate in weed species has been an extremely infrequent event, despite over 20 years of extensive use. Recently, a glyphosate-resistant biotype of goosegrass (Eleusine indica) was identified in Malaysia exhibiting an LD(50) value approximately 2- to 4-fold greater than the sensitive biotype collected from the same region. A comparison of the inhibition of 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) activity by glyphosate in extracts prepared from the resistant (R) and sensitive (S) biotypes revealed an approximately 5-fold higher IC(50)(glyphosate) for the (R) biotype. Sequence comparisons of the predicted EPSPS mature protein coding regions from both biotypes revealed four single-nucleotide differences, two of which result in amino acid changes. One of these changes, a proline to serine substitution at position 106 in the (R) biotype, corresponds to a substitution previously identified in a glyphosate-insensitive EPSPS enzyme from Salmonella typhimurium. Kinetic data generated for the recombinant enzymes suggests that the second substitution identified in the (R) EPSPS does not contribute significantly to its reduced glyphosate sensitivity. Escherichia coli aroA- (EPSPS deficient) strains expressing the mature EPSPS enzyme from the (R) biotype exhibited an approximately 3-fold increase in glyphosate tolerance relative to strains expressing the mature EPSPS from the (S) biotype. These results provide the first evidence for an altered EPSPS enzyme as an underlying component of evolved glyphosate resistance in any plant species.
Matched MeSH terms: Escherichia coli/growth & development
'Mato Peiyu' pomelo (Citrus grandis (L.) Osbeck 'Mato Peiyu') leaves from pruning are currently an agricultural waste. The aim of this study was to isolate essential oils from these leaves through steam distillation (SD) and solvent-free microwave extraction (SFME) and to evaluate their applicability to skin care by analyzing their antimicrobial, antioxidant (diphenyl-1-picrylhydrazyl scavenging assay, β-carotene/linoleic acid assay, and nitric oxide scavenging assay), anti-inflammatory (5-lipoxygenase inhibition assay), and antityrosinase activities. The gas chromatography-mass spectrometry results indicated that the main components of 'Mato Peiyu' leaf essential oils were citronellal and citronellol, with a total percentage of 50.71% and 59.82% for SD and SFME, respectively. The highest bioactivity among all assays was obtained for 5-lipoxygenase inhibition, with an IC50 value of 0.034% (v/v). The MIC90 of the antimicrobial activity of essential oils against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Candida albicans ranged from 0.086% to 0.121% (v/v). Citronellal and citronellol were the main contributors, accounting for at least 54.58% of the essential oil's bioactivity. This paper is the first to report the compositions and bioactivities of 'Mato Peiyu' leaf essential oil, and the results imply that the pomelo leaf essential oil may be applied in skin care.
Matched MeSH terms: Escherichia coli/growth & development