| 1. |
- Govindaraju, Indira, et al.
(author)
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Microscopic and spectroscopic characterization of rice and corn starch
- 2020
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In: Microscopy research and technique (Print). - : John Wiley & Sons. - 1059-910X .- 1097-0029. ; 83:5, s. 490-498
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Journal article (peer-reviewed)abstract
- Starch granules from rice and corn were isolated, and their molecular mechanism on interaction with alpha-amylase was characterized through biochemical test, microscopic imaging, and spectroscopic measurements. The micro-scale structure of starch granules were observed under an optical microscope and their average size was in the range 1-100 mu m. The surface topological structures of starch with micro-holes due to the effect of alpha- amylase were also visualized under scanning electron microscope. The crystallinity was confirmed by X-ray diffraction patterns as well as second-harmonic generation microscopy. The change in chemical bonds before and after hydrolysis of the starch granules by alpha- amylase was determined by Fourier transform infrared spectroscopy. Combination of microscopy and spectroscopy techniques relates structural and chemical features that explain starch enzymatic hydrolysis which will provide a valid basis for future studies in food science and insights into the energy transformation dynamics.
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| 2. |
- Grønberg, Christina, et al.
(author)
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Structure and ion-release mechanism of P IB-4-type ATPases
- 2022
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In: eLife. - : eLife Sciences Publications Ltd. - 2050-084X.
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Journal article (peer-reviewed)abstract
- Abstract Transition metals, such as zinc, are essential micronutrients in all organisms, but alsohighly toxic in excessive amounts. Heavy-metal transporting P-type (PIB) ATPases are crucial forhomeostasis, conferring cellular detoxification and redistribution through transport of these ionsacross cellular membranes. No structural information is available for the PIB-4-ATPases, the subclasswith the broadest cargo scope, and hence even their topology remains elusive. Here, we presentstructures and complementary functional analyses of an archetypal PIB-4-ATPase, sCoaT fromSulfitobacter sp. NAS14-1. The data disclose the architecture, devoid of classical so-called heavy-metal-binding domains (HMBDs), and provide fundamentally new insights into the mechanism anddiversity of heavy-metal transporters. We reveal several novel P-type ATPase features, includinga dual role in heavy-metal release and as an internal counter ion of an invariant histidine. We alsoestablish that the turnover of PIB-ATPases is potassium independent, contrasting to many otherP-type ATPases. Combined with new inhibitory compounds, our results open up for efforts in forexample drug discovery, since PIB-4-ATPases function as virulence factors in many pathogens.
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| 3. |
- Kumar Gahlot, Dharmender, 1985-, et al.
(author)
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Bioengineering of non-pathogenic Escherichia coli to enrich for accumulation of environmental copper
- 2020
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In: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 10
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Journal article (peer-reviewed)abstract
- Heavy metal sequestration from industrial wastes and agricultural soils is a long-standing challenge. This is more critical for copper since copper pollution is hazardous both for the environment and for human health. In this study, we applied an integrated approach of Darwin's theory of natural selection with bacterial genetic engineering to generate a biological system with an application for the accumulation of Cu2+ ions. A library of recombinant non-pathogenic Escherichia coli strains was engineered to express seven potential Cu2+ binding peptides encoded by a 'synthetic degenerate' DNA motif and fused to Maltose Binding Protein (MBP). Most of these peptide-MBP chimeras conferred tolerance to high concentrations of copper sulphate, and in certain cases in the order of 160-fold higher than the recognised EC50 toxic levels of copper in soils. UV-Vis spectroscopic analysis indicated a molar ratio of peptide-copper complexes, while a combination of bioinformatics-based structure modelling, Cu2+ ion docking, and MD simulations of peptide-MBP chimeras corroborated the extent of Cu2+ binding among the peptides. Further, in silico analysis predicted the peptides possessed binding affinity toward a broad range of divalent metal ions. Thus, we report on an efficient, cost-effective, and environment-friendly prototype biological system that is potentially capable of copper bioaccumulation, and which could easily be adapted for the removal of other hazardous heavy metals or the bio-mining of rare metals.
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| 4. |
- Li, Ping, et al.
(author)
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PcoB is a defense outer membrane protein that facilitates cellular uptake of copper
- 2022
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In: Protein Science. - : Wiley. - 0961-8368 .- 1469-896X. ; 31:7
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Journal article (peer-reviewed)abstract
- Copper (Cu) is one of the most abundant trace metals in all organisms, involved in a plethora of cellular processes. Yet elevated concentrations of the element are harmful, and interestingly prokaryotes are more sensitive for environmental Cu stress than humans. Various transport systems are present to maintain intracellular Cu homeostasis, including the prokaryotic plasmid-encoded multiprotein pco operon, which is generally assigned as a defense mechanism against elevated Cu concentrations. Here we structurally and functionally characterize the outer membrane component of the Pco system, PcoB, recovering a 2.0 Å structure, revealing a classical β-barrel architecture. Unexpectedly, we identify a large opening on the extracellular side, linked to a considerably electronegative funnel that becomes narrower towards the periplasm, defining an ion-conducting pathway as also supported by metal binding quantification via inductively coupled plasma mass spectrometry and molecular dynamics (MD) simulations. However, the structure is partially obstructed towards the periplasmic side, and yet flux is permitted in the presence of a Cu gradient as shown by functional characterization in vitro. Complementary in vivo experiments demonstrate that isolated PcoB confers increased sensitivity towards Cu. Aggregated, our findings indicate that PcoB serves to permit Cu import. Thus, it is possible the Pco system physiologically accumulates Cu in the periplasm as a part of an unorthodox defense mechanism against metal stress. These results point to a previously unrecognized principle of maintaining Cu homeostasis and may as such also assist in the understanding and in efforts towards combatting bacterial infections of Pco-harboring pathogens.
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| 5. |
- Mahato, Dhani Ram, et al.
(author)
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Dynamic lipid interactions in the plasma membrane Na+,K+-ATPase
- 2023
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In: Biochimica et Biophysica Acta. Molecular Cell Research. - : Elsevier. - 0167-4889 .- 1879-2596. ; 1870:7
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Journal article (peer-reviewed)abstract
- The function of ion-transporting Na+,K+-ATPases depends on the surrounding lipid environment in biological membranes. Two established lipid-interaction sites A and B within the transmembrane domain have been observed to induce protein activation and stabilization, respectively. In addition, lipid-mediated inhibition has been assigned to a site C, but with the exact location not experimentally confirmed. Also, possible effects on lipid interactions by disease mutants dwelling in the membrane-protein interface remain relatively uncharacterized. We simulated human Na+,K+-ATPase α1β1FXYD homology models in E1 and E2 states in an asymmetric, multicomponent plasma membrane to determine both wild-type and disease mutant lipid-protein interactions. The simulated wild-type lipid interactions at the established sites A and B were in agreement with experimental results thereby confirming the membrane-protein model system. The less well-characterized, proposed inhibitory site C was dominated by lipids lacking inhibitory properties. Instead, two sites hosting inhibitory lipids were identified at the extracellular side and also a cytoplasmic CHL-binding site that provide putative alternative locations of Na+,K+-ATPase inhibition. Three disease mutations, Leu302Arg, Glu840Arg and Met859Arg resided in the lipid-protein interface and caused drastic changes in the lipid interactions. The simulation results show that lipid interactions to the human Na+,K+-ATPase α1β1FXYD protein in the plasma membrane are highly state-dependent and can be disturbed by disease mutations located in the lipid interface, which can open up for new venues to understand genetic disorders.
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| 6. |
- Wiuf, Anders, et al.
(author)
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The two-domain elevator-type mechanism of zinc-transporting ZIP proteins
- 2022
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In: Science Advances. - : American Association for the Advancement of Science. - 2375-2548. ; 8:28
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Journal article (peer-reviewed)abstract
- Zinc is essential for all organisms and yet detrimental at elevated levels. Hence, homeostasis of this metal is tightly regulated. The Zrt/Irt-like proteins (ZIPs) represent the only zinc importers in metazoans. Mutations in human ZIPs cause serious disorders, but the mechanism by which ZIPs transfer zinc remains elusive. Hitherto, structural information is only available for a model member, BbZIP, and as a single, ion-bound conformation, precluding mechanistic insights. Here, we elucidate an inward-open metal-free BbZIP structure, differing substantially in the relative positions of the two separate domains of ZIPs. With accompanying coevolutional analyses, mutagenesis, and uptake assays, the data point to an elevator-type transport mechanism, likely shared within the ZIP family, unifying earlier functional data. Moreover, the structure reveals a previously unknown ninth transmembrane segment that is important for activity in vivo. Our findings outline the mechanistic principles governing ZIP-protein transport and enhance the molecular understanding of ZIP-related disorders.
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