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Natalia P Zakataeva, Dmitriy V Romanenkov, Yuliya R Yusupova, Victoria S Skripnikova, Takayuki Asahara, Sergey V Gronskiy
5'-nucleotidases (EC catalyze the hydrolytic dephosphorylation of 5'-ribonucleotides and 5'-deoxyribonucleotides as well as complex nucleotides, such as uridine 5'-diphosphoglucose (UDP-glucose), nicotinamide adenine dinucleotide and flavin adenine dinucleotide, to their corresponding nucleosides plus phosphate. These enzymes have been found in diverse species in intracellular and membrane-bound, surface-localized forms. Soluble forms of 5'-nucleotidases belong to the ubiquitous haloacid dehalogenase superfamily (HADSF) and have been shown to be involved in the regulation of nucleotide, nucleoside and nicotinamide adenine dinucleotide (NAD+) pools...
2016: PloS One
Shubhangi Kaushik, Zbynek Prokop, Jiri Damborsky, Radka Chaloupkova
Molecular recognition mechanisms and kinetics of binding of ligands to buried active sites via access tunnels are not well understood. Fluorescence polarization enables rapid and non-destructive real-time quantification of the association between small fluorescent ligands and large biomolecules. In this study, we describe analysis of binding kinetics of fluorescent ligands resembling linear halogenated alkanes to haloalkane dehalogenases. Dehalogenases possess buried active sites connected to the surrounding solvent by access tunnels...
November 9, 2016: FEBS Journal
Siyan Zhao, Chang Ding, Jianzhong He
Dehalococcoides mccartyi exhibits versatile capabilities to respire halogenated compounds under anaerobic conditions. In this study, we report the assembly and annotation of the complete genome of a chloroethene dechlorinating D. mccartyi strain 11a5. Bearing a 1,461,973 base-pair chromosome, strain 11a5 distinguishes itself from other D. mccartyi strains by possessing a 5,940 base-pair circular extrachromosomal genetic element which contains a reductive dehalogenase homolog. The whole genome of strain 11a5 harbors 31 putative reductive dehalogenase genes...
November 16, 2016: FEMS Microbiology Ecology
Susan Maleki, Radka Hrudikova, Sergey B Zotchev, Helga Ertesvåg
: The alginate-producing bacterium Pseudomonas fluorescens utilizes the Entner-Doudoroff (ED) and pentose phosphate (PP) pathways to metabolize fructose, since the upper part of its Embden-Meyerhof-Parnas pathway is defective. Our previous study indicated that perturbation of the central carbon metabolism by diminishing glucose-6-phosphate dehydrogenase activity could lead to sugar-phosphate stress when P. fluorescens was cultivated on fructose. In the present study, we demonstrate that PFLU2693, annotated as a haloacid dehalogenase-like enzyme, is a new sugar-phosphate phosphatase now designated Spp, which is able to dephosphorylate a range of phosphate substrates including glucose 6-phosphate and fructose 6-phosphate in vitro The effect of spp overexpression on growth and alginate production was investigated using both wild type and several mutant strains...
November 11, 2016: Applied and Environmental Microbiology
Cresten B Mansfeldt, Gretchen W Heavner, Annette R Rowe, Boris Hayete, Bruce W Church, Ruth E Richardson
The interpretation of high-throughput gene expression data for non-model microorganisms remains obscured because of the high fraction of hypothetical genes and the limited number of methods for the robust inference of gene networks. Therefore, to elucidate gene-gene and gene-condition linkages in the bioremediation-important genus Dehalococcoides, we applied a Bayesian inference strategy called Reverse Engineering/Forward Simulation (REFS™) on transcriptomic data collected from two organohalide-respiring communities containing different Dehalococcoides mccartyi strains: the Cornell University mixed community D2 and the commercially available KB-1® bioaugmentation culture...
2016: PloS One
Koshlan Mayer-Blackwell, Maeva Fincker, Olivia Molenda, Benjamin Callahan, Holly Sewell, Susan Holmes, Elizabeth A Edwards, Alfred M Spormann
Bioremediation of groundwater contaminated with chlorinated aliphatic hydrocarbons such as perchloroethene and trichloroethene can result in the accumulation of the undesirable intermediate vinyl chloride. Such accumulation can either be due to the absence of specific vinyl chloride respiring Dehalococcoides mccartyi or to the inhibition of such strains by the metabolism of other microorganisms. The fitness of vinyl chloride respiring Dehalococcoides mccartyi subpopulations is particularly uncertain in the presence of chloroethene/chloroethane cocontaminant mixtures, which are commonly found in contaminated groundwater...
November 3, 2016: Environmental Science & Technology
Aliyu Adamu, Mohd Shahir Shamsir, Roswanira Abdul Wahab, Sepideh Parvizpour, Fahrul Huyop
Dehalogenases are of high interest due to their potential applications in bioremediation and in synthesis of various industrial products. DehL is an L-2-haloacid dehalogenase (EC that catalyses the cleavage of halide ion from L-2-halocarboxylic acid to produce D-2-hydroxycarboxylic acid. Although DehL utilises the same substrates as the other L-2-haloacid dehalogenases, its deduced amino acid sequence is substantially different (<25%) from those of the rest L-2-haloacid dehalogenases. To date, the 3D structure of DehL is not available...
November 21, 2016: Journal of Biomolecular Structure & Dynamics
Jun Li, Yoel Rodriguez, Chunming Cheng, Lei Zeng, Elaine Y M Wong, Chelsea Y Xu, Ming-Ming Zhou, Pin-Xian Xu
EYA1 is known to be overexpressed in human breast cancer in which Myc protein is also accumulated in association with decreased phospho-T58 levels. We have recently reported that EYA1 functions as a unique protein phosphatase to dephosphorylate Myc at phosphor-T58 to regulate Myc levels. However, it remains unclear whether EYA1-mediated Myc dephosphorylation on T58 1 is a critical function in regulating Myc protein stability in breast cancer. Furthermore, EYA1's substrate specificity has remained elusive. In this study, we have investigated these questions and here we report that depletion of EYA1 using shRNA in breast cancer cells destabilizes Myc protein and increases pT58 levels, leading to an increase in doubling time and impairment of cell cycle progression...
October 17, 2016: Molecular and Cellular Biology
Xi-Hui Xu Qin He, Fan Zhang, Yu-Kai Tai, Yan-Fei Luo, Jian He, Qing Hong, Jian-Dong Jiang, Xin Yan
BACKGROUND: To reduce the fermentation cost for industrialization of chlorothalonil hydrolytic dehalogenase (Chd), agro-industrial wastewater including molasses, corn steep liquor (CSL) and fermentation wastewater were used to substitute the expensive carbon and nitrogen sources and fresh water for lab preparation. RESULTS: The results showed that molasses and CSL could replace 5% carbon source and 100% organic nitrogen source to maintain the same fermentation level...
October 8, 2016: Journal of the Science of Food and Agriculture
Bruna Matturro, Carla Ubaldi, Simona Rossetti
The toxicity of polychlorinated biphenyls (PCB) can be efficiently reduced in contaminated marine sediments through the reductive dechlorination (RD) process lead by anaerobic organohalide bacteria. Although the process has been extensively investigated on PCB-spiked sediments, the knowledge on the identity and metabolic potential of PCB-dechlorinating microorganisms in real contaminated matrix is still limited. Aim of this study was to explore the composition and the dynamics of the microbial communities of the marine sediment collected from one of the largest Sites of National Interest (SIN) in Italy (Mar Piccolo, Taranto) under conditions promoting the PCBs RD...
2016: Frontiers in Microbiology
Manaki Mimura, Rémi Zallot, Thomas Daniel Niehaus, Ghulam Hasnain, Satinder K Gidda, Thuy Nd Nguyen, Erin M Anderson, Robert T Mullen, Greg Brown, Alexander F Yakunin, Valerié de Crécy-Lagard, Jesse F Gregory, Donald R McCarty, Andrew D Hanson
To synthesize the cofactor thiamin diphosphate (ThDP), plants must first hydrolyze thiamin monophosphate (ThMP) to thiamin, but dedicated enzymes for this hydrolysis step were unknown and widely doubted to exist. The classical thiamin-requiring th2-1 mutation in Arabidopsis was shown to reduce ThDP levels by half and to increase ThMP levels five-fold, implying that the THIAMIN REQUIRING 2 (TH2) gene product could be a dedicated ThMP phosphatase. Genomic and transcriptomic data indicated that TH2 corresponds to At5g32470, encoding a HAD (haloacid dehalogenase) family phosphatase fused to a TenA (thiamin salvage) family protein...
September 27, 2016: Plant Cell
Yasutomo Shinohara, Shunji Takahashi, Hiroyuki Osada, Yasuji Koyama
Esterified drimane-type sesquiterpene lactones such as astellolides display various biological activities and are widely produced by plants and fungi. Given their low homology to known sesquiterpene cyclases, the genes responsible for their biosynthesis have not been uncovered yet. Here, we identified the astellolide gene cluster from Aspergillus oryzae and discovered a novel sesquiterpene biosynthetic machinery consisting of AstC, AstI, and AstK. All these enzymes are annotated as haloacid dehalogenase-like hydrolases, whereas AstC also contains a DxDTT motif conserved in class II diterpene cyclases...
2016: Scientific Reports
Lydia Krasper, Hauke Lilie, Anja Kublik, Lorenz Adrian, Ralph Golbik, Ute Lechner
: Reductive dehalogenases are essential enzymes in organohalide respiration and consist of a catalytic subunit A and a membrane protein B, encoded by rdhAB genes. Thirty-two rdhAB genes exist in the genome of Dehalococcoides mccartyi strain CBDB1. To gain a first insight into the regulation of rdh operons, the control of gene expression of two rdhAB genes (cbdbA1453/cbdbA1452 and cbdbA1455/cbdbA1454) by the MarR-type regulator Rdh2R (cbdbA1456) encoded directly upstream was studied using heterologous expression and in vitro studies...
December 1, 2016: Journal of Bacteriology
Shan Shan, Haowei Min, Ting Liu, Dunquan Jiang, Zihe Rao
Trehalose serves as a key structural component in the cell wall of Mycobacterium tuberculosis. M. tuberculosis trehalose-6-phosphate phosphatase (MtbTPP), an essential enzyme in the trehalose biosynthesis OtsAB pathway, catalyzes the dephosphorylation of trehalose-6-phosphate (trehalose-6-P) to generate trehalose, and plays a critical role in M. tuberculosis survival-associated cell wall formation and permeability. Therefore, MtbTPP (OtsB2) is considered a promising potential target for discovery of antimicrobial drugs...
December 2016: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
Yanwei Li, Ruiming Zhang, Likai Du, Qingzhu Zhang, Wenxing Wang
The quantum mechanics/molecular mechanics (QM/MM) method (e.g., density functional theory (DFT)/MM) is important in elucidating enzymatic mechanisms. It is indispensable to study "multiple" conformations of enzymes to get unbiased energetic and structural results. One challenging problem, however, is to determine the minimum number of conformations for DFT/MM calculations. Here, we propose two convergence criteria, namely the Boltzmann-weighted average barrier and the disproportionate effect, to tentatively address this issue...
2016: International Journal of Molecular Sciences
Rui-Juan Niu, Qing-Chuan Zheng, Hong-Xing Zhang
The haloalkaloic acid dehalogenase (HAD) phosphatase from Thermococcus onnurineus NA1 (TON_0338), has phosphatase activity the flavin mono-nucleotide (FMN). The molecular origin and structural motifs for the activity deficiency of double-tryptophan mutant have not been rationalized at atomic resolution. Molecular dynamics (MD) simulations and the molecular mechanics/Generalized-Born surface area (MM/GBSA) free energy calculations were used to explore the effects of mutations on the changes in both structural flexibility and conformational dynamics...
September 2016: Journal of Molecular Graphics & Modelling
Samuel H Schneider, Steven G Boxer
IR and Raman frequency shifts have been reported for numerous probes of enzyme transition states, leading to diverse interpretations. In the case of the model enzyme ketosteroid isomerase (KSI), we have argued that IR spectral shifts for a carbonyl probe at the active site can provide a connection between the active site electric field and the activation free energy (Fried et al. Science 2014, 346, 1510-1514). Here we generalize this approach to a much broader set of carbonyl probes (e.g., oxoesters, thioesters, and amides), first establishing the sensitivity of each probe to an electric field using vibrational Stark spectroscopy, vibrational solvatochromism, and MD simulations, and then applying these results to reinterpret data already in the literature for enzymes such as 4-chlorobenzoyl-CoA dehalogenase and serine proteases...
September 15, 2016: Journal of Physical Chemistry. B
Iren Wang, Szu-Yu Chen, Shang-Te Danny Hsu
DehI is a homodimeric haloacid dehalogenase from Pseudomonas putida that contains the most complex 61 Stevedore's protein knot within its folding topology. To examine how DehI attains such an intricate knotted topology we combined far-UV circular dichroism (CD), intrinsic fluorescence spectroscopy and small angle X-ray scattering (SAXS) to investigate its folding mechanism. Equilibrium unfolding of DehI by chemical denaturation indicated the presence of two highly populated folding intermediates, I and I'. While the two intermediates vary in secondary structure contents and tertiary packing according to CD and intrinsic fluorescence, respectively, their overall dimension and compactness are similar according to SAXS...
2016: Scientific Reports
Anett Schallmey, Marcus Schallmey
Halohydrin dehalogenases are industrially relevant enzymes that catalyze the reversible dehalogenation of vicinal haloalcohols with formation of the corresponding epoxides. In the reverse reaction, also other negatively charged nucleophiles such as azide, cyanide, or nitrite are accepted besides halides to open the epoxide ring. Thus, novel C-N, C-C, or C-O bonds can be formed by halohydrin dehalogenases, which makes them attractive biocatalysts for the production of various β-substituted alcohols. Despite the fact that only five individual halohydrin dehalogenase enzyme sequences have been known until recently enabling their heterologous production, a large number of different biocatalytic applications have been reported using these enzymes...
September 2016: Applied Microbiology and Biotechnology
Na Sa, Renu Rawat, Chelsea Thornburg, Kevin D Walker, Sanja Roje
Despite the importance of riboflavin as the direct precursor of the cofactors flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN), the physiologically relevant catalyst dephosphorylating the riboflavin biosynthesis pathway intermediate 5-amino-6-ribitylamino-2,4(1H,3H) pyrimidinedione 5'-phosphate (ARPP) has not been characterized from any organism. By using as the query sequence a previously identified plastidial FMN hydrolase AtcpFHy1 (At1g79790), belonging to the haloacid dehalogenase (HAD) superfamily, seven candidates for the missing ARPP phosphatase were found, cloned, recombinantly expressed, and purified...
August 4, 2016: Plant Journal: for Cell and Molecular Biology
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