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Purification of Tropomyosin Br-3 and 5NM1 and Characterization of Their Interactions with Actin

Varnyuné Kis-Bicskei, Nikolett and Leipold Vig, Andrea and Nyitrai, Miklós and Bugyi, Beáta and Talián, Csaba Gábor (2013) Purification of Tropomyosin Br-3 and 5NM1 and Characterization of Their Interactions with Actin. CYTOSKELETON, 70 (11). pp. 755-765. ISSN 1949-3584

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Abstract

Tropomyosins were first identified in neuronal systems in 1973. Although numerous isoforms were found and described since then, many aspects of their function and interactions remained unknown. Tropomyosin isoforms show different sorting pattern in neurogenesis. As one example, TM5NM1/2 is present in developing axons, but it is replaced by TMBr-3 in mature neurons, suggesting that these tropomyosin isoforms contribute differently to the establishment of the functional features of the neuronal actin networks. We developed a method for the efficient purification of TMBr-3 and TM5NM1 as recombinant proteins using bacterial expression system and investigated their interactions with actin. We found that both isoforms bind actin filaments, however, the binding of TM5NM1 was much stronger than that of TMBr-3. TMBr-3 and TM5NM1 modestly affected actin assembly kinetics, in an opposite manner. Consistently with the higher affinity of TM5NM1 it inhibited actin filament disassembly more efficiently than TMBr-3. Similarly to other previously studied tropomyosins TM5NM1 inhibited the Arp2/3 complex-mediated actin assembly. Notably, TMBr-3 did not influence the Arp2/3 complex-mediated polymerization. This is a unique feature of TMBr-3, since so far it is the only known tropomyosin supporting the activity of the Arp2/3 complex, indicating that TMBr-3 may colocalize and work simultaneously with Arp2/3 complex in neuronal cells.

Item Type: Article
Uncontrolled Keywords: actin; tropomyosin; actin nucleation factors; purification; fluorescence spectroscopy
Subjects: Q Science / természettudomány > QH Natural history / természetrajz > QH301 Biology / biológia > QH3020 Biophysics / biofizika
SWORD Depositor: MTMT SWORD
Depositing User: MTMT SWORD
Date Deposited: 07 Feb 2014 10:08
Last Modified: 31 Dec 2015 00:15
URI: http://real.mtak.hu/id/eprint/10156

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