Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The plasma and serum of humans and various animal species exert an actin-depolymerizing activity. Human actin-depolymerizing factor (ADF) has been purified by ammonium sulfate fractionation, DEAE-cellulose and blue-Sepharose chromatography. It is a single polypeptide of approximately 90 kDa, with a pI between 6.0 and 6.5. ADF is heat and trypsin-sensitive, inactivated by EGTA, not stained by HIO4/Schiff on sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS/PAGE), and not retained on a concanavalin-A-Sepharose column. Incubation of ethanol-fixed cultured cells or unfixed cryostat tissue sections with ADF abolishes immunofluorescent actin staining, by a mechanism which involves extraction of actin from the preparations. ADF promotes fragmentation and depolymerization of actin filaments as shown by electron microscopy, differential ultracentrifugation and DNAse I inhibition assay. This depolymerized actin retains its mobility on SDS/PAGE and is able to repolymerize in the presence of EGTA. Human white blood cells and platelets (but neither human fibroblasts nor white blood cells and platelets from pig, rat and rabbit) contain a 90-kDa protein reacting with an antibody raised in rabbit against human ADF as judged by immunofluorescence and immunoblotting techniques. Immunoblots of human granulocyte subcellular fractions suggest that the protein reacting with ADF antibody is present in the soluble cytoplasmic fraction. ADF may play a role in solubilization of plasma actin and in the intracellular organization of actin, and should be useful for the evaluation of the relative stability of cytoplasmic actin filaments in various physiological and pathological processes.
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PMID:Human plasma actin-depolymerizing factor. Purification, biological activity and localization in leukocytes and platelets. 257 90

Using separation of total cellular proteins by two dimensional (2-D) gel electrophoresis (isoelectric focusing/SDS-PAGE) we have characterized two regulated proteins, p21 and p19, in dog thyroid cells. We have used the same 2-D gel technique to purify these proteins before their trypsin cleavage and partial sequencing. Three peptides were sequenced in the case of p19 and two peptides in the case of p21. The Swiss-Prot protein sequence database revealed that p19 was identical to destrin/ADF (actin depolymerizing factor) and p21 to cofilin, two closely related and widely distributed actin-binding proteins. This was further verified by cross-reactivity with specific antibodies against brain cofilin and chicken ADF. We have demonstrated, using 2-D gel electrophoresis with a nonequilibrium pH gradient in the first dimension (nonequilibrium pH gradient in the first dimension (nonequilibrium pH gradient electrophoresis/SDS-PAGE) that, in the thyroid cell, cofilin and destrin/ADF were present, under control conditions, in two forms: a phosphorylated and an unphosphorylated one. Thyrotropin (TSH), through cyclic AMP, provoked a very rapid dephosphorylation of these two proteins, which was already maximal after 20 min of action, whereas their dephosphorylation in response to 12-O-tetradecanoylphorbol-13-acetate (TPA) was slower. This suggests that dephosphorylation of cofilin and destrin/ADF by TSH could be implicated in the disruption of actin-containing stress fibers and in the reorganization of microfilaments induced by this hormone. Epidermal growth factor, which does not induce acute morphological changes in thyroid cells, did not affect the state of phosphorylation of cofilin and destrin/ADF except for a delayed decrease (after 24 h) of destrin/ADF phosphorylation. A 10% dimethyl sulfoxide treatment of thyroid cells also induced rapid dephosphorylation of destrin and cofilin. This was accompanied by a reorganization of actin microfilaments that clearly resembles the one induced by TSH and by the appearance of intranuclear cofilin-containing rods. However, these rod structures were not observed in response to TSH, forskolin, or TPA, suggesting that dephosphorylation of cofilin correlates with the reorganization of actin microfilaments but not with the nuclear transport of cofilin. We propose that the dephosphorylation of destrin and cofilin could be involved in the TSH-stimulated macropinocytic activity, a key process in thyroid hormone secretion.
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PMID:Characterization and identification as cofilin and destrin of two thyrotropin- and phorbol ester-regulated phosphoproteins in thyroid cells. 817 42

The amino acid sequences of destrin and cofilin are very similar (84% homology) throughout the entire range of proteins, but they have different functions. In this study, we constructed a new cofilin expression plasmid, which had high expression frequency, and the structures of destrin and cofilin were analyzed by limited proteolysis and circular dichroism (CD). When destrin was digested by trypsin, two fragments of 17.0 kDa and 9.2 kDa were obtained, whereas only one 8.4 kDa fragment was obtained from cofilin. In spite of the overall sequence homology, an N-terminal amino acid sequence analyses of the fragments revealed the cleavage sites on destrin and cofilin to be different. These results suggest that destrin and cofilin differ in their overall tertiary folds. Cofilin showed activity similar to destrin at high pH values, although no pH-dependent structural change in cofilin was confirmed by using limited proteolysis and CD.
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PMID:Evidence for structural differences between the two highly homologous actin-regulatory proteins, destrin and cofilin. 953 78

Destrin is a 19 kDa actin-depolymerizing protein of the ADF-cofilin family. Destrin was digested with trypsin to a structurally stable 9.2 kDa fragment that contains the actin-binding sequence. The purified 9.2 kDa fragment has an actin filament stabilizing activity, rather than an actin filament depolymerizing activity. The deleted region is probably essential for the actin filament depolymerizing activity of intact destrin. Surprisingly, the 9.2 kDa fragment also has an assembly-promoting activity in the absence of ATP.
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PMID:The actin-depolymerizing factor destrin has an actin-stabilizing domain. 1180 18