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The E. coli strain, overproducing human growth hormone (hGH) was made by integration of the hGH gene under the control of T7 promoter into the chromosomal LacZ gene of BL21(DE3) via lambda Red recombineering. The strain gave higher productivity (50 mg·L−1·OD550−1) and better growth characteristics than the corresponding strain in which the same hGH expression cassette was placed in a plasmid. The...
A long standing debate in biochemistry is to determine whether the conformational changes observed during biomolecular interactions proceed through conformational selection (of preexisting isoforms) or induced fit (ligand-induced 3D reshaping). The latter mechanism had been invoked in certain circumstances, for example to explain the non-Michaelian activity of monomeric enzymes like glucokinase. But...
We have proposed an allosteric ATP inhibition mechanism of Pfk-2 determining the structure of different forms of the enzyme together with a kinetic enzyme analysis. Here we complement the mechanism by using hybrid oligomers of the homodimeric enzyme to get insights about the allosteric communication pathways between the same sites or different ones located in different subunits. Kinetic analysis of...
Leucine stimulates anabolic and catabolic processes in skeletal muscle, however little is known about the effects of leucine on peroxisome proliferator-activated receptor (PPAR) activity. This work characterized the effects of 24-h leucine treatment on metabolic parameters and protein expression in cultured myotubes. Leucine significantly increased PPARβ/δ expression as well as markers of mitochondrial...
Ribosomal protein L19 is an essential ribosomal protein and is a component of bridge B8, one of the protein-RNA bridges linking the large and small ribosomal subunits. Bridge B8 also contributes to the accuracy of translation by affecting GTPase activation by ribosome-bound aminoacyl tRNA-EF-Tu•GTP ternary complexes. Previous work has identified a limited number of accuracy-altering alterations in...
Oxidatively damaged DNA bases are substrates for two overlapping repair pathways: DNA glycosylase-initiated base excision repair (BER) and apurinic/apyrimidinic (AP) endonuclease-initiated nucleotide incision repair (NIR). In the BER pathway, an AP endonuclease cleaves DNA at AP sites and 3′-blocking moieties generated by DNA glycosylases, whereas in the NIR pathway, the same AP endonuclease incises...
The tumor suppressor protein p53 is a key factor in genome stability and one of the most studied of DNA binding proteins. This is the first study on the interaction of wild-type p53 with guanine quadruplexes formed by the human telomere sequence. Using electromobility shift assay and ELISA, we show that p53 binding to telomeric G-quadruplexes increases with the number of telomeric repeats. Further,...
ADAMs are transmembrane multifunctional proteins that contain disintegrin and metalloprotease domains. ADAMs act in a diverse set of biological processes, including fertilization, inflammatory responses, myogenesis, cell migration, cell proliferation and ectodomain cleavage of membrane proteins. These proteins also have additional functions in pathological processes as cancer and metastasis development...
The chloride channel (ClC) protein family comprises both chloride (Cl−) channels and chloride/proton (Cl−/H+) antiporters. In prokaryotes and eukaryotes, these proteins mediate the movement of Cl− ions across the membrane. In eukaryotes, ClC proteins play a role in the stabilization of membrane potential, epithelial ion transport, hippocampal neuroprotection, cardiac pacemaker activity and vesicular...
Efforts to identify genes and characterize enzymes involved in the biosynthesis of plant cell wall polysaccharides have yet to produce and purify to homogeneity an active plant cell wall synthesizing enzyme suitable for structural studies. In Arabidopsis, the last step of xyloglucan (XG) biosynthesis is catalyzed by fucosyltransferase 1 (AtFUT1), which transfers l-fucose from GDP-β-l-fucose to a specific...
Loxosceles spiders' venom comprises a complex mixture of biologically active toxins, mostly consisting of low molecular mass components (2–40 kDa). Amongst, isoforms of astacin-like metalloproteases were identified through transcriptome and proteome analyses. Only LALP1 (Loxosceles Astacin-Like protease 1) has been characterized. Herein, we characterized LALP3 as a novel recombinant astacin-like metalloprotease...
Chitinase 3-like 1, otherwise known as YKL-40, is a secreted glycoprotein purported to have a role in extracellular matrix metabolism. The first mammalian cell type found to express YKL-40 was the human osteosarcoma-derived osteoblast, MG63. In that first study the active vitamin D3 metabolite, 1,25-dihydroxycholecalciferol (1,25D), stimulated YKL-40 expression, thereby indicating that a vital factor...
Research using 1,8-naphthalimide derivatives has expanded rapidly in recent years owing to their cell-permeable nature, ability to target certain cellular locations and fluorescent properties. Here we describe the synthesis of three new esters of 4-hydroxy-N-propyl-1,8-naphthalimide (NAP) and the development of a simple and sensitive assay protocol to measure the activity of carboxylester hydrolases...
Antimicrobial activity of thiosulfinates in situ produced by mixtures of Citrobacter freundii methionine γ-lyase (MGL) with new substrates, l-methionine and S-(alkyl/allyl)-l-cysteine sulfoxides has been recently demonstrated (Anufrieva et al., 2015). This opens a way to the rational design of a new biotechnologically relevant antimicrobial drug producer. To increase the efficiency of the enzyme toward...
EcoP15I DNA methyltransferase (M.EcoP15I) recognizes short asymmetric sequence, 5′-CAGCAG-3′, and methylates the second adenine only on one strand of the double-stranded DNA (dsDNA). In vivo, this methylation is sufficient to protect the host DNA from cleavage by the cognate restriction endonuclease, R.EcoP15I, because of the stringent cleavage specificity requirements. Biochemical and structural...
Nano-formulations of medicinal drugs have attracted the interest of many researchers for drug delivery applications. These nano-formulations enhance the properties of conventional drugs and are specific to the targeted delivery site. Dendrimers, polymeric nanoparticles, liposomes, nano-emulsions and micelles are some of the nano-formulations that are gaining prominence in pharmaceutical industry for...
The ribonucleoprotein enzyme telomerase maintains telomeres and is essential for cellular immortality in most cancers. Insight into the telomerase mechanism can be gained from short telomere syndromes, in which mutation of telomerase components manifests in telomere dysfunction. We carried out detailed kinetic analyses and molecular modelling of a disease-associated mutant in the C-terminal extension...
Occurrence of the adhesion/growth-regulatory galectins as family sets the challenge to achieve a complete network analysis. Along this route taken for a well-suited model organism (chicken), we fill the remaining gap to characterize its seventh member known from rat as galectin-related inter-fiber protein (GRIFIN) in the lens. Its single-copy gene is common to vertebrates, with one or more deviations...
Vitamin B12 and omega-3 fatty acids are important nutrients required for neuronal functioning. We have demonstrated the beneficial effects of vitamin B12 and omega-3 fatty acid supplementation on brain neurotrophins and cognition in the first and second generation offspring. However, there is a need to examine if the effects are sustained in the third generation offspring. This study reports the effects...
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