scholarly journals Identification of a 23 kDa protein from maize photoaffinity-labelled with 5-azido-[7-3H]indol-3-ylacetic acid

1995 ◽  
Vol 305 (3) ◽  
pp. 853-857 ◽  
Author(s):  
J Feldwisch ◽  
R Zettl ◽  
N Campos ◽  
K Palme
Keyword(s):  
Manganese Superoxide Dismutase ◽  
Reversed Phase ◽  
Amino Acid Sequences ◽  
Size Exclusion ◽  
Dichlorophenoxyacetic Acid ◽  
Photoaffinity Labelling ◽  
Manganese Superoxide ◽  
2,4 Dichlorophenoxyacetic Acid ◽  
Triton X ◽  
Ylacetic Acid

A 23 kDa protein (p23) was identified in microsomal extracts from maize coleoptiles by photoaffinity labelling with 5-azido-[7-3H]indol-3-ylacetic acid ([3H]N3IAA). Labelling of p23 was blocked by unlabelled IAA, N3IAA, indol-3-ylbutyric acid and indol-3-yl-lactate. In addition, labelling was efficiently decreased by tryptophan, as well as by the scavenger p-aminobenzoic acid. Labelling was, however, not affected by synthetic auxins such as 1-naphthylacetic acid or 2,4-dichlorophenoxyacetic acid. Competition data suggest that the label was probably bound via the indole ring, and hence labelling was not specific for auxins. The 23 kDa protein was solubilized from crude microsomes by extraction with Triton X-100 and purified to homogeneity by ion-exchange, size-exclusion and reversed-phase chromatography. After electroblotting, the amino acid sequences of the p23 N-terminus as well as the several tryptic peptides were obtained. Database comparisons revealed sequence identity with a maize manganese superoxide dismutase. We conclude that photoaffinity labelling of p23 was pseudo-affinity, and therefore the binding site for IAA is not specific.

Molecular cloning and characterization of the copper/zinc and manganese superoxide dismutase genes from the human parasiteClonorchis sinensis

Parasitology ◽  
2005 ◽  
Vol 130 (6) ◽  
pp. 687-697 ◽  
Author(s):  
A. H. LI ◽  
Y. KONG ◽  
S. H. CHO ◽  
H. W. LEE ◽  
B. K. NA ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Manganese Superoxide Dismutase ◽  
Mrna Level ◽  
Clonorchis Sinensis ◽  
Amino Acid Sequences ◽  
Sod Activity ◽  
Intronless Gene ◽  
Copper Zinc Superoxide Dismutase ◽  
Manganese Superoxide ◽  
Copper Zinc

A copper/zinc superoxide dismutase (Cu/ZnSOD) gene and a manganese superoxide dismutase (MnSOD) gene of the human parasiteClonorchis sinensishave been cloned and their gene products functionally characterized. GenesCu/ZnSODandMnSODencode proteins of 16 kDa and 25·4 kDa, respectively. The deduced amino acid sequences of the two genes contained highly conserved residues required for activity and secondary structure formation of Cu/ZnSOD and MnSOD, respectively, and show up to 73·7% and 75·4% identities with their counterparts in other animals. The genomic DNA sequence analysis of Cu/ZnSOD gene revealed this as an intronless gene. Inhibitor studies with purified recombinant Cu/ZnSOD and MnSOD, both of which were functionally expressed inEscherichia coli, confirmed that they are copper/zinc and manganese-containing SOD, respectively. Immunoblots showed that bothC. sinensisCu/ZnSOD and MnSOD should be antigenic for humans, and both, especially theC. sinensisMnSOD, exhibit extensive cross-reactions with sera of patients infected by other trematodes or cestodes. RT-PCR and SOD activity staining of parasite lysates indicate that there are no significant differences in mRNA level or SOD activity for both species of SOD, indicating cytosolic Cu/ZnSOD and MnSOD might play a comparatively important role in theC. sinensisantioxidant system.

scholarly journals Characteristics of a Novel Manganese Superoxide Dismutase of a Hadal Sea Cucumber (Paelopatides sp.) from the Mariana Trench

Marine Drugs ◽  
2019 ◽  
Vol 17 (2) ◽  
pp. 84 ◽  
Author(s):  
Yanan Li ◽  
Xue Kong ◽  
Haibin Zhang
Keyword(s):  
Superoxide Dismutase ◽  
Metal Ion ◽  
Sea Cucumber ◽  
Manganese Superoxide Dismutase ◽  
Guanidine Hydrochloride ◽  
Tween 20 ◽  
Potential Candidate ◽  
Manganese Superoxide ◽  
Divalent Metal Ion ◽  
Triton X

A novel, cold-adapted, and acid-base stable manganese superoxide dismutase (Ps-Mn-SOD) was cloned from hadal sea cucumber Paelopatides sp. The dimeric recombinant enzyme exhibited approximately 60 kDa in molecular weight, expressed activity from 0 °C to 70 °C with an optimal temperature of 0 °C, and resisted wide pH values from 2.2–13.0 with optimal activity (> 70%) at pH 5.0–12.0. The Km and Vmax of Ps-Mn-SOD were 0.0329 ± 0.0040 mM and 9112 ± 248 U/mg, respectively. At tested conditions, Ps-Mn-SOD was relatively stable in divalent metal ion and other chemicals, such as β-mercaptoethanol, dithiothreitol, Tween 20, Triton X-100, and Chaps. Furthermore, the enzyme showed striking stability in 5 M urea or 4 M guanidine hydrochloride, resisted digestion by proteases, and tolerated a high hydrostatic pressure of 100 MPa. The resistance of Ps-Mn-SOD against low temperature, extreme acidity and alkalinity, chemicals, proteases, and high pressure make it a potential candidate in biopharmaceutical and nutraceutical fields.

scholarly journals Novel 2,4-Dichlorophenoxyacetic Acid Degradation Genes from Oligotrophic Bradyrhizobium sp. Strain HW13 Isolated from a Pristine Environment

2002 ◽  
Vol 184 (2) ◽  
pp. 509-518 ◽  
Author(s):  
Wataru Kitagawa ◽  
Sachiko Takami ◽  
Keisuke Miyauchi ◽  
Eiji Masai ◽  
Yoichi Kamagata ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Burkholderia Cepacia ◽  
Ralstonia Eutropha ◽  
Rhodococcus Erythropolis ◽  
Amino Acid Sequences ◽  
Dichlorophenoxyacetic Acid ◽  
Phenol Derivatives ◽  
Degrading Bacteria ◽  
2,4 Dichlorophenoxyacetic Acid

ABSTRACT The tfd genes of Ralstonia eutropha JMP134 are the only well-characterized set of genes responsible for 2,4-dichlorophenoxyacetic acid (2,4-D) degradation among 2,4-D-degrading bacteria. A new family of 2,4-D degradation genes, cadRABKC, was cloned and characterized from Bradyrhizobium sp. strain HW13, a strain that was isolated from a buried Hawaiian soil that has never experienced anthropogenic chemicals. The cadR gene was inferred to encode an AraC/XylS type of transcriptional regulator from its deduced amino acid sequence. The cadABC genes were predicted to encode 2,4-D oxygenase subunits from their deduced amino acid sequences that showed 46, 44, and 37% identities with the TftA and TftB subunits of 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) oxygenase of Burkholderia cepacia AC1100 and with a putative ferredoxin, ThcC, of Rhodococcus erythropolis NI86/21, respectively. They are thoroughly different from the 2,4-D dioxygenase gene, tfdA, of R. eutropha JMP134. The cadK gene was presumed to encode a 2,4-D transport protein from its deduced amino acid sequence that showed 60% identity with the 2,4-D transporter, TfdK, of strain JMP134. Sinorhizobium meliloti Rm1021 cells containing cadRABKC transformed several phenoxyacetic acids, including 2,4-D and 2,4,5-T, to corresponding phenol derivatives. Frameshift mutations indicated that each of the cadRABC genes was essential for 2,4-D conversion in strain Rm1021 but that cadK was not. Five 2,4-D degraders, including Bradyrhizobium and Sphingomonas strains, were found to have cadA gene homologs, suggesting that these 2,4-D degraders share 2,4-D degradation genes similar to those of strain HW13 cadABC.

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