Diethylpyrocarbonate modification reveals HisB5 as an important modulator of insulin amyloid formation
This study not only provides a fast new approach for studying the impact of imidazole ring in amyloid formation, but also reveals the critical modulating role of histidine imidazole ring on the amyloidogenicity of insulin. (Source: Journal of Biochemistry)
Source: Journal of Biochemistry - January 5, 2015 Category: Biochemistry Authors: Yang, X., Li, Y., Huang, L., Zhang, X., Cheng, C., Gong, H., Ma, L., Huang, K. Tags: Regular Papers Source Type: research
Molecular characterization of acidic peptide:N-glycanase from the dimorphic yeast Yarrowia lipolytica
In this study, we identified and characterized a PNGase A-like enzyme, PNGase Yl, in the dimorphic yeast Yarrowia lipolytica. The corresponding gene was cloned and recombinantly expressed in Pichia pastoris. The purified enzyme cleaved glycans from glycopeptides with the maximum activity at pH 5. No metal ions were required for full activity, and rather it was repressed by three metal ions (Fe3+, Cu2+ and Zn2+). Using glycopeptide substrates, PNGase Yl was shown to release various types of N-glycans including high-mannose and complex-type glycans as well as glycans containing core-linked α(1,3)-fucose that are freque...
Source: Journal of Biochemistry - January 5, 2015 Category: Biochemistry Authors: Lee, K. J., Gil, J. Y., Kim, S.-Y., Kwon, O., Ko, K., Kim, D.-I., Kim, D. K., Kim, H. H., Oh, D.-B. Tags: Regular Papers Source Type: research
The cytoplasmic peptide:N-glycanase (Ngly1)--basic science encounters a human genetic disorder
Peptide:N-glycanase (PNGase) is a de-N-glycosylating enzyme that cleaves intact N-glycans from glycoproteins/glycopeptides. The activity of the cytoplasmic PNGase in several mammalian-derived cultured cells was first reported in 1993, and 7 years later, the gene encoding the enzyme was identified in budding yeast. Although the gene—PNG1 in budding yeast and NGLY1/Ngly1 in mammalian cells—appears to be well conserved throughout eukaryotes, the biological significance of this enzyme has remained elusive until recently. However, discovery of a human genetic disorder involving the NGLY1 gene clearly indicates that ...
Source: Journal of Biochemistry - January 5, 2015 Category: Biochemistry Authors: Suzuki, T. Tags: JB Special Reviews-Fundamental Roles of Glycans in Eukaryotes Source Type: research
Mechanisms for modulation of neural plasticity and axon regeneration by chondroitin sulphate
Chondroitin sulphate proteoglycans (CSPGs), consisting of core proteins linked to one or more chondroitin sulphate (CS) chains, are major extracellular matrix (ECM) components of the central nervous system (CNS). Multi-functionality of CSPGs can be explained by the diversity in structure of CS chains that undergo dynamic changes during development and under pathological conditions. CSPGs, together with other ECM components, form mesh-like structures called perineuronal nets around a subset of neurons. Enzymatic digestion or genetic manipulation of CSPGs reactivates neural plasticity in the adult brain and improves regenera...
Source: Journal of Biochemistry - January 5, 2015 Category: Biochemistry Authors: Miyata, S., Kitagawa, H. Tags: JB Special Reviews-Fundamental Roles of Glycans in Eukaryotes Source Type: research
Glycobiology of {alpha}-dystroglycan and muscular dystrophy
Most proteins are modified by glycans, which can modulate the biological properties and functions of glycoproteins. The major glycans can be classified into N-glycans and O-glycans according to their glycan-peptide linkage. This review will provide an overview of the O-mannosyl glycans, one subtype of O-glycans. Originally, O-mannosyl glycan was only known to be present on a limited number of glycoproteins, especially α-dystroglycan (α-DG). However, once a clear relationship was established between O-mannosyl glycan and the pathological mechanisms of some congenital muscular dystrophies in humans, research on t...
Source: Journal of Biochemistry - January 5, 2015 Category: Biochemistry Authors: Endo, T. Tags: JB Special Reviews-Fundamental Roles of Glycans in Eukaryotes Source Type: research
Conformational stability, reversibility and heat-induced aggregation of {alpha}-1-acid glycoprotein
To investigate the relationship between conformational stability, reversibility of denaturation and aggregation of protein, we determined the conformation, melting temperature (Tm), and reversibility of heat-induced denaturation of α-1-acid glycoprotein (AGP) in aqueous solutions at various pH values using circular dichroism (CD) and differential scanning microcalorimetry. To quantitate and characterize heat-induced AGP aggregation under the same pH conditions, solutions of AGP were incubated at 50°C and then analysed by size exclusion chromatography (SEC), sodium dodecyl sulfate-polyacrylamide gel electrophoresi...
Source: Journal of Biochemistry - November 26, 2014 Category: Biochemistry Authors: Iwura, T., Fukuda, J., Yamazaki, K., Arisaka, F. Tags: Regular Papers Source Type: research
Identification of the critical residues for the function of vacuolar H+-pyrophosphatase by mutational analysis based on the 3D structure
H+-translocating pyrophosphatase (H+-PPase) converts energy from hydrolysis of inorganic pyrophosphate (PPi) to active H+ translocation across membranes. From the 3D structure resolved by crystallography, 17 amino acid residues in several domains of mung bean (Vigna radiata) enzyme were selected and substituted with alanine individually. The mutant enzymes were expressed in yeast cells to evaluate their biochemical role. The highly conserved residues in the substrate-binding site (T249, D269, D507 and N534) were shown to be essential for PPi hydrolysis and H+ pump. The amino acid substitution of residues in the H+ transloc...
Source: Journal of Biochemistry - November 26, 2014 Category: Biochemistry Authors: Asaoka, M., Segami, S., Maeshima, M. Tags: Regular Papers Source Type: research
DNA-rescuable allosteric inhibition of aptamer II ligand affinity by aptamer I element in the shortened Vibrio cholerae glycine riboswitch
Glycine riboswitches contain two aptamers and turn on the expression of downstream genes in bacteria. Although full-length glycine riboswitches were shown to exhibit no glycine-binding cooperativity, the truncated glycine riboswitches were confirmed to bind two glycine molecules cooperatively. Thorough understanding of the ligand-binding cooperativity may shed light on the molecular basis of the cooperativity and help design novel intricate biosensing genetic circuits for application in synthetic biology. A previously proposed sequential model does not readily provide explanation for published data showing a deleterious mu...
Source: Journal of Biochemistry - November 26, 2014 Category: Biochemistry Authors: Sherman, E. M., Elsayed, G., Esquiaqui, J. M., Elsayed, M., Brinda, B., Ye, J.-D. Tags: Regular Papers Source Type: research
Identification of 14,20-dihydroxy-docosahexaenoic acid as a novel anti-inflammatory metabolite
Docosahexaenoic acid (DHA) exhibits anti-inflammatory activity related to some of its oxygenated metabolites, such as D-series resolvins, protectin and maresin. Here, we analysed the lipids in inflammatory exudates using liquid chromatography–tandem mass spectrometry and identified a novel DHA metabolite, 14,20-dihydroxy-DHA (14,20-diHDHA) and showed that it is biosynthesized by eosinophils through the 12/15-lipoxygenase pathway. The chemical structure of the dominant 14,20-diHDHA isomer, which is endogenously biosynthesized by eosinophils, was identified as 14S,20R-diHDHA using chemically synthesized stereoisomers. ...
Source: Journal of Biochemistry - November 26, 2014 Category: Biochemistry Authors: Yokokura, Y., Isobe, Y., Matsueda, S., Iwamoto, R., Goto, T., Yoshioka, T., Urabe, D., Inoue, M., Arai, H., Arita, M. Tags: Regular Papers Source Type: research
Nuclear import of human histone lysine-specific demethylase LSD1
Upregulation and nuclear retention of the human histone demethylase LSD1 are correlated with aggressiveness and poor outcome of several cancer types, but the molecular mechanism of LSD1 nuclear import remains unclear. Here, we found that the N-terminal flexible region of LSD1 contains a nuclear localization signal (NLS), 112RRKRAK117. Mutation or deletion of the NLS completely abolished the nuclear import of LSD1, suggesting the motif is a bona fide NLS. More importantly, our GST pull-down assay showed that LSD1 physically interacts with three proteins of importin α family. In addition, our data suggest that the nucl...
Source: Journal of Biochemistry - November 26, 2014 Category: Biochemistry Authors: Jin, Y., Kim, T. Y., Kim, M. S., Kim, M. A., Park, S. H., Jang, Y. K. Tags: Regular Papers Source Type: research
The Gateway Reflex, which is mediated by the inflammation amplifier, directs pathogenic immune cells into the CNS
The brain–blood barrier (BBB) tightly limits immune cell migration into the central nervous system (CNS), avoiding unwanted inflammation under the normal state. However, immune cells can traverse the BBB when inflammation occurs within the CNS, suggesting a certain signal that creates a gateway that bypasses the BBB might exist. We revealed the inflammation amplifier as a mechanism of this signal, and identified dorsal vessels of the fifth lumber (L5) spinal cord as the gateway. The inflammation amplifier is driven by a simultaneous activation of NF-B and STATs in non-immune cells, causing the production of a large a...
Source: Journal of Biochemistry - November 26, 2014 Category: Biochemistry Authors: Sabharwal, L., Kamimura, D., Meng, J., Bando, H., Ogura, H., Nakayama, C., Jiang, J.-J., Kumai, N., Suzuki, H., Atsumi, T., Arima, Y., Murakami, M. Tags: JB Review Source Type: research
Effect of non-native helix destabilization on the folding of equine {beta}-lactoglobulin
β-lactoglobulin forms a non-native α-helix during an early stage of folding. To address the role of the non-native structure in the folding process, we designed several mutants of equine β-lactoglobulin with reduced helical propensity in the non-native helix region. One of them, A123T, showed a similar structure to that of the wild-type protein; its folding kinetics was investigated by stopped-flow circular dichroism (CD) and fluorescence. Although A123T showed a reduced burst-phase CD intensity, its folding rate was similar to that of the wild-type protein, which indicated that the formation of the non-nat...
Source: Journal of Biochemistry - October 27, 2014 Category: Biochemistry Authors: Okabe, T., Miyajima, T., Nakagawa, K., Tsukamoto, S., Fujiwara, K., Ikeguchi, M. Tags: Regular Papers Source Type: research
Molecular mechanisms for the p38-induced cellular senescence in normal human fibroblast
In this study, we identified the MAPK p38 as a downstream mediator of TAK1, which represses hTERT transcription. Further, we observed that hTERT expression was repressed in senescent normal human fibroblast, and was attenuated on treatment with SB203580, a p38-specific inhibitor, which suggests that p38 represses hTERT expression during cellular senescence. Next, we demonstrated that repression of hTERT, irrespective of the activation status of p38, is important for the induction of cellular senescence, by using hTERT-overexpressing cells and hTERT-knockdown cells. Our results suggested that p38 is activated during the ser...
Source: Journal of Biochemistry - October 27, 2014 Category: Biochemistry Authors: Harada, G., Neng, Q., Fujiki, T., Katakura, Y. Tags: Regular Papers Source Type: research
Macromolecular crowding decelerates aggregation of a {beta}-rich protein, bovine carbonic anhydrase: a case study
In this study, we investigated the effect of synthetic macromolecular crowders on bovine carbonic anhydrase (BCA, a β-rich protein) aggregation. In contrast to the effect of macromolecular crowding on α-rich proteins, BCA aggregation was observed to be reduced due to decrease in the population of aggregation-prone intermediates as a consequence of increased native state stability. In addition, the extent of aggregation was found to depend on the nature of the crowder under consideration. Combining the published data on α-proteins and this study, we conclude that macromolecular crowding can have opposite co...
Source: Journal of Biochemistry - October 27, 2014 Category: Biochemistry Authors: Mittal, S., Singh, L. R. Tags: Regular Papers Source Type: research
Interaction of the muscarinic acetylcholine receptor M2 subtype with G protein G{alpha}i/o isotypes and G{beta}{gamma} subunits as studied with the maltose-binding protein-M2-G{alpha}i/o fusion proteins expressed in Escherichia coli
We expressed the fusion proteins of the muscarinic acetylcholine receptor M2 subtype (M2 receptor) with a maltose-binding protein (MBP) and various G protein α subunits (Gαi1–i3/o) at its N- and C-terminals, respectively (MBP-M2-Gαi/o), in Escherichia coli, and examined the effect of G protein β subunits (Gβ) on the receptor–Gα interaction as assessed by agonist- and GDP-dependent [35S]GTPS binding of the fusion proteins. We found that (i) Gβ promoted both the agonist-dependent and -independent [35S]GTPS binding with little effect on the guanine nucleotide-sensitive high-a...
Source: Journal of Biochemistry - October 27, 2014 Category: Biochemistry Authors: Ichiyama, S., Nemoto, R., Tanabe, H., Haga, T. Tags: Regular Papers Source Type: research
