PubMed Health⌕ Search

Biomedical subjects

E W Daoud

Publications and source records attributed to E W Daoud.

6 recordsLinked to original sources
PubMed1988

Properties of purified actin depolymerizing factor from chick brain.

Actin depolymerizing factor (ADF) from 19-day embryonic chick brains has been purified to greater than 98% homogeneity with a yield of 7.2 mg/100 g of brain. Quantitative immunoblotting with a monospecific antibody to ADF indicated that ADF comprises 0.3% of the total brain protein, resulting in an actual purification yield of about 20%. Brain ADF migrates as a single polypeptide of 19,000 kDa on SDS-containing polyacrylamide gels. The molecular weight of the native protein determined from sedimentation equilibrium in buffers containing from 50 to 200 mM KCl is 20,000. The secondary structure of ADF calculated from the circular dichroic spectrum consists of about 22% alpha-helix, 24% beta-sheet, and 18% beta-turn. ADF contains a blocked N-terminus, a single tryptophan residue located about one-third of the way from one end of the protein, and six cysteine residues (all in reduced form in the native protein). All six cysteine residues could be chemically modified with eosinylmaleimide under nondenaturing conditions; however, ADF activity was lost when more than one cysteine residue was modified. ADF microheterogeneity has been observed upon nonequilibrium pH gradient electrophoresis in polyacrylamide gels containing 9 M urea, the major isoform having a pI of congruent to 7.9-8.0. ADF can interact with either monomeric or filamentous actin to give a complex which can be isolated by gel filtration chromatography. Both major and minor isoforms of the ADF are found in the complex. Assembly-competent actin and active ADF can both be recovered from the complex by chromatography on ATP-saturated DEAE-cellulose.(ABSTRACT TRUNCATED AT 250 WORDS)

Actin Depolymerizing Factors↗
PubMed1988

Inhibition of deoxyribonuclease I activity by actin covalently cross-linked to chick brain actin depolymerizing factor through exposed sulfhydryls.

All but one of the six free sulfhydryl groups of chick brain actin depolymerizing factor (ADF) are protected from modification when ADF forms a 1:1 complex with actin. This exposed sulfhydryl can be cross-linked to cys 374 of actin with N,N'-phenylenedimaleimide. The cross-linked complex inhibits the hydrolytic activity of pancreatic deoxyribonuclease (DNase I) to an identical extent as both the untreated complex and an equivalent amount of free actin. These data indicate that ADF binds to actin at a site which does not overlap with the DNase I binding site.

Actin Depolymerizing Factors↗
PubMed1987

Purification and characterization of phosphodiesterase (exonuclease) from Cerastes cerastes (Egyptian sand viper) venom.

A venom exonuclease 'phosphodiesterase' (E.C. 3.1.4.1) has been purified from Cerastes cerastes venom by a combination of gel filtration on Sephadex G-100 superfine and ion exchange chromatography on DEAE-Sepharose. The enzyme showed a single band on PAGE and SDS-PAGE and had a molecular weight of 110,000. The final preparation was purified 28 fold. It had no carbohydrate and it did not have protease or 5'-nucleotidase activities. Optimum temperature for enzyme activity was 56 degrees C. The enzyme was rapidly inactivated when pre-incubated above 40 degrees C. Energy of activation (Ea) was calculated to be 0.913. The optimum pH was 9.0. Cysteine, glutathione, dithiothreitol, 2-mercaptoethanol, ADP and AMP inhibited the enzyme. Cysteine caused a non-competitive inhibition, while ADP showed a competitive inhibition. EDTA at a concentration of 0.5 mM caused complete inhibition of the enzyme, which could be reversed by the addition of Ca2+ or Mn2+.

Animals↗
PubMed1987

Further characterization of the anticoagulant proteinase, cerastase F-4 from Cerastes cerastes (Egyptian sand viper) venom.

A potent anticoagulant, cerastase F-4, was purified from the venom of Cerastes cerastes. The u.v. absorption spectrum revealed a relatively high tyrosine and low tryptophan content. The molar extension coefficient and E278(0.1%) were 19,400 and 0.84, respectively. The enzyme secondary structure, as studied by circular dichroism, showed 23.6% alpha-helix, 34% beta-sheets, 19% beta-turns and 32.5% random coils. When casein was used as a substrate the optimum pH was 10.0 and the Km was 1.45 g/l. Cerastase F-4 is a metallo-enzyme that contains one mole of Ca2+ and one mole of Zn2+ per mole of protein. It is not affected by phenylmethane sulfonylfluoride or soybean trypsin inhibitor, while it is completely inhibited by 0.5 mM EDTA or ethyleneglycol bis (beta-amino ethylether) N,N,N',N'-tetraacetic acid (EGTA). Ca2+, Mg2+ and Zn2+ partially activated the enzyme under different experimental conditions. Our results suggest that Ca2+ and Zn2+ may play a role in maintaining the structural and catalytic integrity of the enzyme.

Animals↗