tailieunhanh - Báo cáo khoa học: Regulation of the actin–myosin interaction by titin

Titin is known to interact with actin thin filaments within the I-band region of striated muscle sarcomeres. In this study, we have used a titin fragment of 800 kDa (T800) purified from striated skeletal muscle to measure the effect of this interaction on the functional properties of the actin– myosin complex. MALDI-TOF MS revealed that T800 contains the entire titin PEVK (Pro, Glu, Val, Lys-rich) 1 domain. In the presence of tropomyosin–troponin, T800 increased the sliding velocity (both average and maximum values) of actin filaments on heavy-meromyosin (HMM)-coated surfaces and dramatically decreased the number of stationary filaments. . | Eur. J. Biochem. 271 4572-4581 2004 FEBS 2004 doi Regulation of the actin-myosin interaction by titin Nicolas Niederlander1 Fabrice Raynaud2 Catherine Astier2 and Patrick Chaussepied1 1CRBM-CNRS Montpellier France 2EPHE-UMR5539-CNRS Montpellier France Titin is known to interact with actin thin filaments within the I-band region of striated muscle sarcomeres. In this study we have used a titin fragment of 800 kDa T800 purified from striated skeletal muscle to measure the effect of this interaction on the functional properties of the actinmyosin complex. MALDI-TOF MS revealed that T800 contains the entire titin PEVK Pro Glu Val Lys-rich domain. In the presence of tropomyosin-troponin T800 increased the sliding velocity both average and maximum values of actin filaments on heavy-meromyosin HMM -coated surfaces and dramatically decreased the number of stationary filaments. These results were correlated with a 30 reduction in actin-activated HMM ATPase activity and with an inhibition of HMM binding to actin N-terminal residues as shown by chemical cross-linking. At the same time T800 did not affect the efficiency of the Ca2 -controlled on off switch nor did it alter the overall binding energetics of HMM to actin as revealed by cosedimentation experiments. These data are consistent with a competitive effect of PEVK domain-containing T800 on the electrostatic contacts at the actin-HMM interface. They also suggest that titin may participate in the regulation of the active tension generated by the actin-myosin complex. Keywords ATPase chemical cross-linking mass spectrometry motility assay muscle contraction. Titin is the largest known protein containing more than 38 000 residues in its longest human striated muscle isoform. It represents the third most abundant component of vertebrate striated muscle after myosin and actin and is also present in smooth muscle and nonmuscle cells recently reviewed in 1 2 . The importance of intact titin for