From Proteopedia

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F-Actin

Filamentous actin (F-actin) units are also referred to as microfilament ^[1] and are highly conserved, proteinous components found near ubiquitously in eukaryotic cytoskeletons. F-actin and other actin proteins generally have structural roles in cells.

Introduction

Actin is found in nearly all eukaryotic cells and is known primarily for its function as a structural and translocation protein. It also has an ATPase function, as it hydrolyzes ATP to ADP and Pi and undergoes conformational changes with each hydrolysis. Actin belongs to the actin superfamily, which includes other proteins such as Hsp70(DnaK), Hsc70, and hexokinase, because of its nucelotide-dependent conformational change^[2]. Because of the similarity observed in Escherichia Coli's, Hsc70 and ATPase domain of actin, it is believed that the two proteins have a common ancestory^[3]. Prokaryotes are not known to have actin, but do however have an actin homologue, MreB, which also leads to the idea of possible common ancestory^[4].

Actin occurs in two forms: globular actin (G-actin), the free monomeric units of actin, and filamentous actin (F-actin) which is the polymer form. These two forms exist in a dynamic equilibrium with one another as ATP-associated polymerization and depolymerization occur continuously within the cell.

Assembly

G-actin is the free monomeric form of actin which transitions to F-actin. The structures of globular and filamentous actin are distinct from one another in numerous ways, despite the fact that G-actin comprises F-actin. When the monomeric actin becomes polymerized into F-actin, the unit becomes flattened. G-actin appears to have more ion ligands in its structure, and also has the ligand RHO as opposed to 4-methyl histidine as found in the F-actin structure.

Formation of F-actin is a dynamic process of assembly and disassembly which has been termed “treadmilling”. The transition between G- and F-actin begins with a stabilized oligomer of ATP-actin units formed through a nucleation-condensation type fold pattern^[5]. Addition of ATP-monomeric units to either end subsequently occurs, however, because of a difference in charge polarity in the two ends, there is preferential addition to what is termed the "plus (+) end" or the "barbed-end". On the opposite end, the "minus (-) end" or the "pointed end", there is preferential dissociation of actin units^[6]. After attachment of the ATP-bound actin, hydrolysis of the ATP occurs yielding the ADP+Pi bound state. Subsequent loss of a Pi leaves the ADP-actin state^[7]. Because of the potential for addition or removal of monomeric units to occur at both ends, the assembly of F-actin may be described in terms of equilibrium. However, because the rate of ATP-actin association is ten-fold that of ADP-actin dissociation, the f-actin has the appearance of moving forward, or "treadmilling"^[8]. ADP-actin monomers dissociate at the minus end and become recycled to ATP-actin so polymerization at the plus end may occur once again.

Structure

History of the structure

The F-actin protein was discovered by Straub in 1942. The structure was speculated based on a low-resolution x-ray crystallograph found in 1990 by Holmes et al. and over this time, despite the importance of F-actin in eukaryotic cells, this speculated structure was accepted. A higher resolution structure was only recently deposited in the PDB databank in Decemeber 2008 by Oda et al. ^[9].

Domains of F-actin Unit

Structure of a unit of F-actin with domains from a single polypeptide chain. Note the cleft between the two domains houses the nucleotide phosphate ligand and the Ca2+ metal ion ligand. Domain movement is made possible by rotation about the , shown in purple. According to Oda et al.^[10]Domain 2 is believed to tilt 20 degrees and fit itself with Domain 1, thus giving a flatter conformation than the free G-actin. It is not certain whether this flattening occurs before or after ATP hydrolysis.

Stability

The flattened folded form of F-actin requires different stabilization mechanisms than the free monomeric G-actin form. Stability of the f-actin complex is achieved by a series of salt bridge formations involving arginine, glutamate, aspartate, and HIC73, a charged methylated histidine residue. Additional stability is believed to arise from interactions across subunits with residues Leu110 and Thr194^[11].

Active Site

The cleavage of the gamma-phosphoryl from the bound ATP is a result of a conformational change upon binding that moves the Gln137 residue closer to the ATP-Ca2+ ligand. Release of the inorganic phosphate occurs via the conformational change of the flexible "D-loop" into an ordered alpha-helix^[12].

Polymer F-actin

F-actin has the appearance of two right-handed helices, with a gradual twist around one another. It is actually composed of repeats of 13 actin units for every 6 left-handed turns, spanning a length of 350 Å. ^[13]. Including the ADP and Ca²⁺, the F-actin molecule as shown here consists of 377 residues (43kDa), two major domains separated by a nucleotide-binding cleft^[14]. Depending on the state of the bound nucleotide, the most stable conformation of F-actin changes. In its ATP and ADP + Pi nucleotide bound states, it has a closed binding cleft. In its ADP only bound state, it has a wider binding cleft^[15]. A characteristic trait of actin is that the domains remain twisted relative to one another, despite the nucleotide-state-dependent conformational changes^[16].

Function

F-actin performs a structural, mechanical, and enzymatic role within eukaryotic cells. These functions are not necessarily distinct.

Cytoskeleton

F-actin is the most abundant component of the cytoskeleton of eukaryotes. It provides large amounts of tensile strength, considering its thin size. In cases where the flexibility is not desirable as a strucutral component, crosslinkages can be formed between F-actin polymers to give greater stiffness and support^[17].

Elongation of F-actin branches leads to the phenomenon of pushing of the plasma membrane forward in lamellopodial and filopodial extension^[18]. This process relies on the dynamic equlibrium state in which G- and F-actin exist, as it is the continual polymerization of actin units on the leading edge that propels the membrane extension. Without the enzymatic ATPase function of F-actin, this process would not be possible.

Actin-Myosin

The relatively flatter shape of F-actin as compared to G-actin means that myosin preferentially binds to F-actin and not G-actin. This means that F-actin is the functional form of actin composing a large part of the thin filaments that function in muscle contraction^[19]. Additionally, the structure of F-actin gives it large resistance to extensive forces, such as those experienced in muscle contraction^[20].

References

1. ↑ Microfilaments - Wikipedia, the free encyclopedia. http://en.wikipedia.org/wiki/Microfilaments. Date accessed: March 16th, 2010.
2. ↑ Graceffa
3. ↑ Holmes1
4. ↑ holmes2
5. ↑ pfaendtner
6. ↑ mitchinson
7. ↑ chen
8. ↑ clasier
9. ↑ Oda T, Iwasa M, Aihara T, Maéda Y, and Narita A. 2009. The nature of the globular-to fibrous actin transition. Nature,457(7228):441-445. PMID: 19158791
10. ↑ oda
11. ↑ oda
12. ↑ graceffa
13. ↑ Holmes, K.C., Popp, D., Gebhard, W. and Kabsch, W. 1990. Atomic model of the actin filament. Nature,347(6288):44-49. PMID: 2395461
14. ↑ oda
15. ↑ pfaendtner
16. ↑ oda
17. ↑ mitchinson
18. ↑ chen
19. ↑ Holmes 2, Holmes et al 3
20. ↑ Mitchinson