Control of Power Generation in Actin-Myosin Gels

Yamamoto Sho

Way of Control over Power Generation in Actin-Myosin gels

３、 2Ca exchange used dialysis membrane

４、 Photolysis of caged

2Ca

１、 Temperature adjustment in the sample generation２、 ATP regeneration system used PC , CPK

１、 Temperature adjustment in the sample generation

・ All the preparations and implements were kept to be cold. （ about at 0 ℃ ）

・ When observing,I put the samples on the THERMO PLATE which was set at 3 ℃

Result

At room temperature(24℃) On THERMO PLATE

Problem ・ I could not control the exact temperature of the “actin-myosin gels”.・ The probe particles swayed from one side to another.(Probably because of the watere current in the THERMO PLATE.)

Solution・ To cover a box to keep the inner temperature cold .

・ Not to use oil immersion lens.( The oil conduct heat of the objective lens to the samples.)

exchange used dialysis membrane2Ca 3.

Jan Scrimgeour,*ab Jae Kyu Cho,c Victor Breedveldc and Jennifer Curtisab,Soft Matter,2011

ProblemPrepolymer solution did not be polymerized.Polyethylene glycol diacrylate has MEHQ as inhibitor.

Solution

Distillation under reduced pressure Boiling point of MEHQ : 243 ℃ 760mmHg 126 ℃ 11mmHg

5.Cooling water 9.Vacuum pump

2Ca

In the cell , force generation on actin-myosin network is controlled by calcium ion concentration. After released by endoplasmic reticulum , calcium is combined with troponin , and tropomyosin is caused a structural change , and finally actin is made activated.

Function of in Muscular Tissue

Method・ F-actin and Tn-Tm complex are mixed to final concentrations of 1mg/ml each in the presence of 0.1M KCl , 0.1mM ATP , 10mM-TrisHCl(pH=8.0) and the mixture are heated at 45 ℃ for 15 min and then cool at a slow rate (about 0.3 deg/min) to room temperature.

To confirm F-actin is crosslinked with TN-TM , I should observe superprecipitation in the presence of Ca ion.

Hajime Honda7 and Sho Asakura,1988

(a) and (c) Video-recorded images of fluorescent actin filaments and (b) and (d) changes in position of the actin filaments brought about by their movement in 0.6 s at (a) and (b) pCa = 3, (c) and (d) pCa = 5.8 at 30℃.

Hajime Honda7 and Sho Asakura,1988

Hajime Honda7 and Sho Asakura,1988

The average verocities at pCa=3 of regulated actin(continuous line with filled circle)and unregulated actin(broken line with open triangles).The broken line with open circle shows the temperature-dependence of regulated actin-activated myosin ATPase activity.