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Beneficial Lactic Acid Bacteria
Published in K. Balamurugan, U. Prithika, Pocket Guide to Bacterial Infections, 2019
Restriction fragment length polymorphism (RFLP) is characterized by the use of restriction enzymes to digest DNA and following separation of the restriction fragments according to their length by agarose gel electrophoresis (Adzitey et al. 2013). Single digestion with AciI of rpoB gene (coding for RNA polymerase β-subunit) in Leuconostoc, Oenococcus, Pediococcus, and two or three digestions (AciI, HinfI and MseI) in Lactobacillus spp. allowed to identify LAB species commonly isolated from wine (Claisse et al. 2007). Restriction patterns of the tuf gene, encoding the elongation factor Tu and universally distributed in gram-positive bacteria, derived by enzymes AluI and HaeIII could effectively differentiate closely related Lactobacillus species (Park et al. 2012). However, sometimes strain variations could not be demonstrated by the RFLP analysis. Morphologic differences (colony shape and size) were evident between Lactobacillus kefir strains ATCC 35411 and ATCC 8007, but genotypic results failed to differentiate them (Mainville et al. 2006).
Three-Dimensional Structure of p21 and Its Implications
Published in Juan Carlos Lacal, Frank McCormick, The ras Superfamily of GTPases, 2017
New guanine nucleotide-binding (GNB) proteins keep on being discovered continuously and the number of individual members of this family of proteins may be 50 to 100 for a higher organism. In order to understand their function and mechanism of action, it is important to get a detailed description of the three-dimensional structure of these proteins down to the atomic level. This has been achieved for p21, the product of the ras protooncogene. It has enabled us to understand the basic features of this molecule whose general function is as a molecular switch. Although the three-dimensional structure of no other GNB protein except bacterial elongation factor Tu (EF-Tu) has been determined, albeit at lower resolution, the results of the structure determination have implications for other GNB proteins as well.
Replicase
Published in Paul Pumpens, Single-Stranded RNA Phages, 2020
The elongation factor Tu-Ts complex, when covalently crosslinked with dimethyl suberimidate, was able to reconstitute functional Qβ replicase, although it lacked the ability to catalyze the known host functions catalyzed by the individual elongation factors (Brown and Blumenthal 1976b). The sample of the Qβ replicase with crosslinked EF-TuTs replacing the individual elongation factors lacked EF-Tu and EF-Ts activities but could initiate transcription of both polyC and Qβ RNA normally and had approximately the same specific activity as the control enzyme. Remarkably, the denatured Qβ replicase formed with the crosslinked EF-TuTs was found to renature much more rapidly than untreated enzyme and, in contrast to the normal replicase, its renaturation was not inhibited by GDP. Moreover, the aminoacyl-tRNA binding site of EF-Tu was not required for the Qβ replicase activity, since the latter was not affected by inhibitors of the aminoacyl-tRNA binding activity of EF-Tu (Brown and Blumenthal 1976a). Thus, when Qβ replicase was treated with kirromycin, an antibiotic which modified EF-Tu activity, the Qβ RNA replication activity was only slightly affected, while the protein synthetic activity of the EF-Tu in the replicase complex was eliminated. The treatment of the pure EF-Tu with kirromycin, however, prevented it from functioning in the renaturation of the Qβ replicase (Brown and Blumenthal 1976a). Remarkably, kirromycin induced a similar conformational change in EF-Tu as the EF-Ts did, thereby opening the guanine nucleotide binding site, and increased tenfold the trypsin cleavage rate of the EF-Tu (Blumenthal et al. 1977). The trypsin-cleaved EF-Tu still could bind GDP and EF-Ts and could function in the Qβ replicase, but it was no longer spontaneously renatured following denaturation in urea.
The microbiome of deep-sea fish reveals new microbial species and a sparsity of antibiotic resistance genes
Published in Gut Microbes, 2021
Fergus W. J. Collins, Calum J. Walsh, Beatriz Gomez-Sala, Elena Guijarro-García, David Stokes, Klara B. Jakobsdóttir, Kristján Kristjánsson, Finlay Burns, Paul D. Cotter, Mary C. Rea, Colin Hill, R. Paul Ross
Bowtie2 was used to align the paired-end metagenomic reads from the deep-sea dataset against the MEGARes database in order to determine the abundance of known antibiotic resistance genes in the samples.30 Antibiotic resistance genes were found in less than half of all samples analyzed (Table 2). The resistance profile of these samples is dominated by resistance to a class of antibiotics known as elfamycins, which target elongation factor TU (EF-Tu) in bacterial cells.31 Potential resistance to elfamycins was identified in 20 of the metagenomic samples analyzed, in each case due to mutations in the EF-Tu encoding gene tufA. As most bacteria encode two virtually identical copies of the EF-Tu genes, levels of actual resistance due to differences in these genes may be overrepresented.32 As elfamycins are not used therapeutically, the mutations identified here may simply be a result of natural variation in the tufA genes rather than resistance due to the exposure to these antibiotics. EF-Tu has been identified as possibly having an important role in maintaining protein synthesis in response to high-pressure treatment of bacteria, thus potential adaptions of these genes to this environment may confer resistance to these antibiotics.33
Proteomics of Pseudomonas aeruginosa: the increasing role of post-translational modifications
Published in Expert Review of Proteomics, 2018
Charlotte Gaviard, Thierry Jouenne, Julie Hardouin
Quantitative proteomic studies, using data-dependent acquisition (DDA), have been performed in different bacteria [80,84–90] and have highlighted differences in the abundance of modified peptides across samples. Isotope labeling and multiplexing methods seem to be one of the method of choice to quantify PTMs. For example, Kosono et al. [84] have observed the upregulation of acetylated sites on 40 proteins (acetate kinase, protein kinase PrkA or sporulation protein) and succinylated sites on 42 proteins (elongation factor Tu, ribosomal protein or enolase) by using a quantitative SILAC approach in B. subtilis in different carbon sources. Stable isotope-based quantification and phosphostaining 2D gel was recently used to highlight the impact of H2O2 stress on phosphoproteome of Bacillus pumilus [91].
Low Abundance of Lactococcus lactis in Human Colorectal Cancer Is Associated with Decreased Natural Killer Cells
Published in Nutrition and Cancer, 2022
Huan Li, Xinhao Du, Li Yan, Zhenzhen Tang, Ling Zhang, Qiao Zheng, Xianghao Zeng, Guimei Chen, Huawen Yue, Xiangsheng Fu
FISH results in the present study were further validated in 20 samples by L. lactis-specific PCR primers designed. The housekeeping gene Tuf, encoding for the elongation factor Tu, was chosen as target for the design of L. lactis specific primers. The sequences were listed as following: Tuf2 forward (5′-TGA ACC ACA ATG GGT TGC TA-3′) and Tuf2 reverse (5′TCG ACT GGA AGA AGG AGT GG-3′).