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Literature summary for 5.3.4.1 extracted from

  • Barnewitz, K.; Guo, C.; Sevvana, M.; Ma, Q.; Sheldrick, G.M.; S๖ling, H.D.; Ferrari, D.M.
    Mapping of a substrate binding site in the protein disulfide isomerase-related chaperone Wind based on protein function and crystal structure (2004), J. Biol. Chem., 279, 39829-39837.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
DNA sequence determination, expression of GFP-tagged wild-type enzyme and mutants Y53S and K84D in COS-7 cells, expression of C-terminally or N-terminally His6-tagged wild-type and mutant Wind in Escherichia coli strain XL 1-Blue and in Vero cells, the affinity for substrate Pipe is higher with the N-terminally His-tagged enzyme compared to the C-terminally tagged one Drosophila melanogaster

Protein Variants

Protein Variants Comment Organism
C24S site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
C27S site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
D29N site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
D31N site-directed mutagenesis, unstable protein that still dimerizes but then aggregates, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
D31N/R41S site-directed mutagenesis, monomeric mutant, slightly decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
D50A site-directed mutagenesis, inactive mutant, normal dimerization Drosophila melanogaster
D50N site-directed mutagenesis, decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
D50S site-directed mutagenesis, altered Pipe targeting compared to the wild-type enzyme, normal dimerization Drosophila melanogaster
D85N site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
E32K site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
E60A site-directed mutagenesis, unaltered Pipe processing and targeting efficiency compared to the wild-type enzyme Drosophila melanogaster
E60Q site-directed mutagenesis, decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
E60Y site-directed mutagenesis, unaltered Pipe processing and targeting efficiency compared to the wild-type enzyme Drosophila melanogaster
E88K site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
E88Q site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
E90R site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
E90R/D29N site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
E90R/E60A site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
G87S site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
H59Y site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
K58S site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
K84D site-directed mutagenesis, decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
K84N site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
K84S site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
L219S/E212Q site-directed mutagenesis, decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
L232K site-directed mutagenesis, increased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
P106D site-directed mutagenesis, similar to the wild-type enzyme, altered conformation Drosophila melanogaster
R215A site-directed mutagenesis, unaltered Pipe processing efficiency but eliminated Pipe targeting compared to the wild-type enzyme Drosophila melanogaster
R218D site-directed mutagenesis, increased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
R41S site-directed mutagenesis, sligtly reduced dimerization of the mutant and processing of Pipe Drosophila melanogaster
T25K site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
T63K site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
V28D site-directed mutagenesis, monomeric mutant, no dimerization, mutant aggregates Drosophila melanogaster
V28Y site-directed mutagenesis, monomeric mutant, no dimerization, mutant aggregates Drosophila melanogaster
Y53S site-directed mutagenesis, decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
Y55K site-directed mutagenesis, mutant shows increased affinity for substrate Pipe but decreased processing efficiency compared to the wild-type enzyme Drosophila melanogaster
Y55K/D31N/R41S site-directed mutagenesis, slightly decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
Y55S site-directed mutagenesis, decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
Y86F site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
Y86L site-directed mutagenesis, highly decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
Y86Q site-directed mutagenesis, highly decreased Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster
Y86S site-directed mutagenesis, unaltered Pipe processing efficiency compared to the wild-type enzyme Drosophila melanogaster

Organism

Organism UniProt Comment Textmining
Drosophila melanogaster
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-
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Purification (Commentary)

Purification (Comment) Organism
recombinant His6-tagged wild-type and mutant Wind from Escherichia coli strain XL 1-Blue by nickel affinity chromatography Drosophila melanogaster

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
Pipe processing and targeting of Pipe, Pipe is an essential Golgi transmembrane-O-sulfotransferase, protein disulfide isomerase-related chaperone Wind is required for processing and correct targeting of the substrate, mapping of multiple substrate binding sites in Pipe, one enzyme site in vicinity of an exposed cluster of tyrosine residues within the thioredoxin fold domain is essential for activity, a second enzyme site in the enzyme's D-domain is also necessary for processing activity, but competitive to the thioredoxin fold domain residue, overview Drosophila melanogaster ?
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?

Subunits

Subunits Comment Organism
dimer
-
Drosophila melanogaster
More protein disulfide isomerase-related chaperone Wind contains a thioredoxin fold domain and a C-terminal D-domain unique in PDI-D proteins Drosophila melanogaster

Synonyms

Synonyms Comment Organism
PDI
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Drosophila melanogaster
protein disulfide isomerase
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Drosophila melanogaster
protein disulfide isomerase-related chaperone Wind
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Drosophila melanogaster