Estimation of denaturation of actin in the actin-myosin complex treated with various conditions by DNAase I inhibition assay.
1. Experiments were conducted to evaluate whether DNAase I (EC 3.1.4.5) inhibition assay was a valuable tool to study the denaturation of actin in the actin-myosin complex treated with various conditions. 2. A sample containing F-actin or natural actomyosin(myosin B) was treated with KI-ATP solution to convert a form which inhibits DNAase I as effectively as G-actin, and the total amount of native actin was determined by DNAase I inhibition assay. 3. On the basis of the values for remaining native actin in the sample obtained by this assay, a percentage of denaturation of actin during treatment was calculated. 4. The present result demonstrated that DNAase I inhibition assay was easy to perform, very sensitive (0.5-2.0 microgram actin) and highly specific for estimating denaturation of actin in the actin-myosin complex treated with heat or high salt concentrations. 5. In addition, the use of DNAase I and standard G-actin preparations stored frozen at -80 degrees C for the assay was found to be possible within a fixed period of time (about 2 weeks), which was helpful in monitoring the denaturation process of actin treated under various conditions for a long period.
Ikeuchi
Y
Y
Laboratory of Biochemistry and Technology of Animal Products, Faculty of Agriculture, University of Niigata, Japan.
Iwamura
K
K
Suzuki
A
A
D000199:Actins / Q000737:chemistry*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D002138:Calibration; D002623:Chemistry Techniques, Analytical / Q000379:methods; D003850:Deoxyribonuclease I / Q000037:antagonists & inhibitors*; D005069:Evaluation Studies as Topic; D007700:Kinetics; D009218:Myosins / Q000737:chemistry*; D011489:Protein Denaturation; D011817:Rabbits; D012492:Salts; D013696:Temperature
D000199:Actins; D012492:Salts; D003850:Deoxyribonuclease I; D000251:Adenosine Triphosphatases; D009218:Myosins