Group:MUZIC:actinin2: Difference between revisions

Muscle cells are responsible for the voluntary and involuntary contraction. Their ultra-structure contains myofibrils, which are bundles mainly formed by actin and myosin filaments. These filaments are organized into repetitive subunits, called sarcomeres, which are connected in tandem by Z-disks, constituting an intricate macromolecular assembly. A plethora of proteins have been identified in the Z-disk [[http://www.proteopedia.org/wiki/index.php/Group:MUZIC:Interactome]], members of different protein classes - globular, intrinsically disordered and multi-domain proteins, but the details about their structure and interaction network at molecular level are still an enigma. Understanding how these proteins work together and how they interact with other molecules can have major impacts in medicine.

The protein α-actinin skeletal muscle isoform 2 (ACTN2) is one the molecule that has a pivotal role in the formation and integrity of the ultra-structure of striated muscle Z-disk. Including, in the beginning steps of progression of  the [[Z-bodies]] in premyofibrils and nascent myofibrils to Z-disks of the mature  [[myofibrils]] <ref>PMID: 16465476</ref>.

ACTN2 is a calcium-independent isoform expressed in all muscle fibers. The expression in human skeletal muscle overlaps ACTN3. Both proteins form heterodimers in vitro and in vivo and share high protein sequence identity (80%),  therefore suggesting a high 3-D structure similarity. They are regulated by phosphoinositides, rendering alpha-actinin capable of binding to titin (and some other Z-disk partners). It is worth pointing out that the other α-actinin non-muscles isoforms (1 and 4) are calcium-dependent. ACTN1 is associate with cell adhesion molecules, stabilizing cell adhesion and regulating cell shape and cell motility, while ACTN4 are related to the cytoskeleton playing an essential role in cell motility and shape and tumor suppressor activity.

ACTN2 [http://www.uniprot.org/uniprot/P35609 UNIPROT (Alpha-actinin-2)] protein is composed of 894 residues organized in a modular fashion (Scheme 1). At the N-terminus it is identified two calponin homology domains (CH), also called actin biding domain(ABD), followed by four spectrin-like repeats domains (R1-4 domain) - referred as rod domain. Finally, the C-terminus is composed of a CaM-like domain (EFhands 1-4). The quaternary structure of ACTN2 is constituted of a stable antiparellel homodimer (~200 kDa) that cross-links anti-parallel actin  filaments and interacts with titin[[http://www.proteopedia.org/wiki/index.php/Group:MUZIC:Titin]] mediated by ABD domain and EF-hand domain, respectively (Scheme 2).

 

 

 

 

[[Image:actinin2_sequence_annotation.jpg]]

 

Scheme 2 - Proposed scheme of the antiparallel actinin dimer structure in the closed and opened conformation after addition of PIP2 (Adapted from Young and Gautel, 2000).  

 

 

The skeletal isoform 2 and 3 are Ca+2-independent isoform protein that seems to be regulate by phosphatidylinositol 4, 5 biphosphate (PIP2). The molecular details of this interaction are not well understood. But, there are evidences that the polar head of PIP2 interact on the ABD domain triggering in one hand a conformation change in this domain to expose more the actin binding sites, increasing the binding interface for F-actin cross-linking and in other hand releasing the EFhands domains to interact to titin.

There is no high resolution structure available for the entire alpha-actinin protein isoforms (ACTN1-4). However, a gallery of structure of alpha-actinin fragment are available (see the link)[[http://www.proteopedia.org/wiki/index.php/Actinin#3D_Structures_of_Actinin]]. The X-ray structure of the ABD domain reveals the arrangement of the two calponin homology domains (CH1 and CH2) in a closed conformation<ref>PMID:15808860</ref>  

The crystal structure of rod domain shows the four spectrins like domain of one chain forms a homo anti-parallel dimer, which is a left handed twisted from one end of the rod to the other end <ref>PMID:11470434</ref>.  

The solution structure of the complex between the human ACTN2 EF-hand domain and the chicken titin Z-repeat 7 domain was solved by NMR spectroscopy. A specific set of contacts and some important residues for protein-protein interactions were identified. It was shown that recognition and positioning of Z-repeat in cavity of EF-hand domain is mediated by balance of electrostatic and hydrophobic interactions. The reported semi-open conformation of complex is commonly seen in homologous structures. This conformation seems to be typical for calcium-independent recognition by EF-hand domain and is fully compatible with formation of stable long-term complexes between titin and α-actinin.

<Structure load='1H8B ' size='200' frame='true' align='right' caption='Fig. 3 - ACTN2 EF-hand domain and the chicken titin Z-repeat 7 domain was solved by NMR spectroscopy [[1h8b]]' scene='insert optional scene name here' /> <scene name='User:Euripides_Ribeiro/workbench/alpha-actinin/Efhand34_zr7titin/2'>(Fig. 3)</scene>

<Structure load='1HCI' size='200' frame='true' align='right' caption='Fig. 2 - Crystal structure of rod domain of human skeletal muscle alpha-actinin [[1hci]]' scene='insert optional scene name here' /> <scene name='User:Euripides_Ribeiro/workbench/alpha-actinin/Rod_domain_sr_dimer/3'>(Fig. 2)</scene>

<Structure load='1TJT' size='200' frame='true' align='right' caption='Fig. 1- Actinin binding domain of actinin-3 human isoform [[1tjt]]' scene='Insert optional scene name here' /> <scene name='User:Euripides_Ribeiro/workbench/alpha-actinin/Actin_binding_domain/11'>(Fig. 1)</scene>.

Hypertrophic cardiomyopathy (HCM) is a genetically transmitted disease and an important cause of morbidity and sudden cardiac death in young people, including competitive athletes <ref>PMID: 7641357</ref>. Recently, Chiu et al (2010) <ref>PMID: 20022194</ref> have reported a list of point mutation in the ''ACTN2'' gene related to hypertrophic cardiomyopathy (Gln9Arg, Gly111Val, Ala119Thr, Thr495Met, Glu583Ala, Glu628Gly, Arg759Thr). However, the impact of these mutation in the ACTN2 protein structure and function is poorly understood.