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101.
We investigated the molecular bases for resistance to several classes of herbicides that bind tubulins in green foxtail (Setaria viridis L. Beauv.). We identified two alpha- and two beta-tubulin genes in green foxtail. Sequence comparison between resistant and sensitive plants revealed two mutations, a leucine-to-phenylalanine change at position 136 and a threonine-to-isoleucine change at position 239, in the gene encoding alpha2-tubulin. Association of mutation at position 239 with herbicide resistance was demonstrated using near-isogenic lines derived from interspecific pairings between green foxtail and foxtail millet (Setaria italica L. Beauv.), and herbicide sensitivity bioassays combined with allele-specific PCR-mediated genotyping. Association of mutation at position 136 with herbicide resistance was demonstrated using herbicide sensitivity bioassays combined with allele-specific PCR-mediated genotyping. Both mutations were associated with recessive cross resistance to dinitroanilines and benzoic acids, no change in sensitivity to benzamides, and hypersensitivity to carbamates. Using three-dimensional modeling, we found that the two mutations are adjacent and located into a region involved in tubulin dimer-dimer contact. Comparison of three-dimensional alpha-tubulin models for organisms with contrasted sensitivity to tubulin-binding herbicides enabled us to propose that residue 253 and the vicinity of the side chain of residue 251 are critical determinants for the differences in herbicide sensitivity observed between organisms, and that positions 16, 24, 136, 239, 252, and 268 are involved in modulating sensitivity to these herbicides. 相似文献
102.
van der Sanden MH Meems H Houweling M Helms JB Vaandrager AB 《The Journal of biological chemistry》2004,279(50):52007-52015
103.
104.
Barindra Sana Ke Ding Jia Wei Siau Rupali Reddy Pasula Sharon Chee Sharad Kharel Jean-Baptise Henri Lena Eunice Goh Lakshminarayanan Rajamani Yeng Ming Lam Sierin Lim John F. Ghadessy 《Biotechnology and bioengineering》2023,120(11):3200-3209
Polyethylene terephthalate (PET) hydrolase enzymes show promise for enzymatic PET degradation and green recycling of single-use PET vessels representing a major source of global pollution. Their full potential can be unlocked with enzyme engineering to render activities on recalcitrant PET substrates commensurate with cost-effective recycling at scale. Thermostability is a highly desirable property in industrial enzymes, often imparting increased robustness and significantly reducing quantities required. To date, most engineered PET hydrolases show improved thermostability over their parental enzymes. Here, we report engineered thermostable variants of Ideonella sakaiensis PET hydrolase enzyme (IsPETase) developed using two scaffolding strategies. The first employed SpyCatcher-SpyTag technology to covalently cyclize IsPETase, resulting in increased thermostability that was concomitant with reduced turnover of PET substrates compared to native IsPETase. The second approach using a GFP-nanobody fusion protein (vGFP) as a scaffold yielded a construct with a melting temperature of 80°C. This was further increased to 85°C when a thermostable PETase variant (FAST PETase) was scaffolded into vGFP, the highest reported so far for an engineered PET hydrolase derived from IsPETase. Thermostability enhancement using the vGFP scaffold did not compromise activity on PET compared to IsPETase. These contrasting results highlight potential topological and dynamic constraints imposed by scaffold choice as determinants of enzyme activity. 相似文献
105.
McElroy SJ Hobbs S Kallen M Tejera N Rosen MJ Grishin A Matta P Schneider C Upperman J Ford H Polk DB Weitkamp JH 《PloS one》2012,7(5):e38373
Necrotizing enterocolitis (NEC) is the leading cause of gastrointestinal morbidity and mortality in preterm infants. NEC is characterized by an exaggerated inflammatory response to bacterial flora leading to bowel necrosis. Bacterial lipopolysaccharide (LPS) mediates inflammation through TLR4 activation and is a key molecule in the pathogenesis of NEC. However, LPS also induces cyclooxygenase-2 (COX-2), which promotes intestinal barrier restitution through stimulation of intestinal cell survival, proliferation, and migration. Epidermal growth factor receptor (EGFR) activation prevents experimental NEC and may play a critical role in LPS-stimulated COX-2 production. We hypothesized that EGFR is required for LPS induction of COX-2 expression. Our data show that inhibiting EGFR kinase activity blocks LPS-induced COX-2 expression in small intestinal epithelial cells. LPS induction of COX-2 requires Src-family kinase signaling while LPS transactivation of EGFR requires matrix metalloprotease (MMP) activity. EGFR tyrosine kinase inhibitors block LPS stimulation of mitogen-activated protein kinase ERK, suggesting an important role of the MAPK/ERK pathway in EGFR-mediated COX-2 expression. LPS stimulates proliferation of IEC-6 cells, but this stimulation is inhibited with either the EGFR kinase inhibitor AG1478, or the selective COX-2 inhibitor Celecoxib. Taken together, these data show that EGFR plays an important role in LPS-induction of COX-2 expression in enterocytes, which may be one mechanism for EGF in inhibition of NEC. 相似文献
106.
Henri Debray Danuta Dus Jean-Michel Wieruszeski Gérard Strecker Jean Montreuil 《Glycoconjugate journal》1991,8(1):29-37
Four bi-antennary glycan fractions of theN-acetyllactosamine-type, derived from a Lewis lung carcinoma (LL2) cell subline resistant to theAleuria aurantia agglutinin were studied by 400 MHz1H-NMR spectroscopy. By this method, their antennae were found to be terminated either by (2-3 or 6)-linkedN-acetylneuraminic acid or (1-3)-linked galactose residues. The primary structure of glycans of these four glycopeptide or derived oligosaccharide-alditols has been determined in full detail.Abbreviations NAc
N-acetyl group
- NGc
N-glycolyl group
- GlcNAc
N-acetylglucosamine
- NeuAc
N-acetylneuraminic acid
- NeuGc
N-glycolylneuraminic acid
- Man
mannose
- Gal
galactose
- Fuc
fucose
- Con A
concanavalin A
- LCA
Lens culinaris agglutinin
- AAA
Aleuria aurantia agglutinin
- WGA
Wheat germ agglutinin
- RCA II
Ricinus communis agglutinin II
- PBS
phosphate buffered saline, 0.01m Na2HPO4/0.14m NaCl, pH 7.2
- HPLC
high performance liquid chromatography
- EMEM
Eagle's Minimal Essential Medium
- LecR
lectin resistant
- MG
-methylglycoside 相似文献
107.
108.
109.
In search of key enzymes in Plasmodium phospholipid metabolism, we demonstrate the presence of a parasite-encoded phosphatidylserine decarboxylase (PSD) in the membrane fraction of Plasmodium falciparum-infected erythrocytes. PSD cDNA, encoding phosphatidylserine decarboxylase (PfPSD), was cloned by screening a directional cDNA library derived from the trophozoite erythrocytic stage. The corresponding PfPSD gene is located on chromosome 9 of P. falciparum, contains one intron of 938 nucleotides and is transcribed into a 3.7 kb mRNA. PfPSD cDNA encodes a putative protein of 362 amino acids, with a predicted molecular mass of 42.6 kDa, which clearly belongs to the type I PSD family. Only a 35 kDa polypeptide was detected in the parasite using a specific rabbit antiserum. PfPSD has a 314VGSS317 sequence near its carboxyl-terminus that is related to the Escherichia coli, yeast and human LGST motif, which is the site of proenzyme processing. PSD enzyme was expressed in E. coli with a KM of 63 +/- 19 microM and a VMAX of 680 +/- 49 nmol of phosphatidylethanolamine formed h-1 mg-1 protein. Site-directed mutagenesis of the VGSS active site demonstrated that the PfPSD proenzyme was processed into two non-identical subunits (alpha and beta) and revealed the crucial role played by each residue in enzyme processing and activity. Using indirect immunofluorescence, PfPSD labelling was co-localized with an endoplasmic reticulum marker, but not with a mitochondrial vital dye. This P. falciparum PSD is the first type I PSD identified in the endoplasmic reticulum compartment. 相似文献
110.
Broët P Lewin A Richardson S Dalmasso C Magdelenat H 《Bioinformatics (Oxford, England)》2004,20(16):2562-2571
MOTIVATION: Multiclass response (MCR) experiments are those in which there are more than two classes to be compared. In these experiments, though the null hypothesis is simple, there are typically many patterns of gene expression changes across the different classes that led to complex alternatives. In this paper, we propose a new strategy for selecting genes in MCR that is based on a flexible mixture model for the marginal distribution of a modified F-statistic. Using this model, false positive and negative discovery rates can be estimated and combined to produce a rule for selecting a subset of genes. Moreover, the method proposed allows calculation of these rates for any predefined subset of genes. RESULTS: We illustrate the performance our approach using simulated datasets and a real breast cancer microarray dataset. In this latter study, we investigate predefined subset of genes and point out interesting differences between three distinct biological pathways. AVAILABILITY: http://www.bgx.org.uk/software.html 相似文献