Structural Basis of Actin Filament Nucleation by Tandem W Domains

Abstract

SUMMARY Spontaneous nucleation of actin is very inefficient in cells. To overcome this barrier, cells have evolved a set of actin filament nucleators to promote rapid nucleation and polymerization in response to specific stimuli. However, the molecular mechanism of actin nucleation remains poorly understood. This is hindered largely by the fact that actin nucleus, once formed, rapidly polymerizes into filament, thus making it impossible to capture stable multisubunit actin nucleus. Here, we report an effective double-mutant strategy to stabilize actin nucleus by preventing further polymerization. Employing this strategy, we solved the crystal structure of AMPPNP-actin in complex with the first two tandem W domains of Cordon-bleu (Cobl), a potent actin filament nucleator. Further sequence comparison and functional studies suggest that the nucleation mechanism of Cobl is probably shared by the p53 cofactor JMY, but not Spire. Moreover, the double-mutant strategy opens the way for atomic mechanistic study of actin nucleation and polymerization.

Mechanisms of leiomodin 2-mediated regulation of actin filament in muscle cells

“…To enable crystallographic studies of biological actin complexes, our group recently has developed a double-mutant strategy in which actin-binding proteins and two types of nonpolymerizable actin mutants are combined to form stable complexes amenable to crystallization (26). This strategy made possible the rapid determination of the first two crystal structures of oligomeric actin with tandem-Gactin-binding nucleator complexes: a mammalian nucleator Cordonbleu (Cobl) (26) and a bacterial effector Vibrio parahaemolyticus protein L (VopL) (28).…”

“…Indeed, although Arp2/3 has been subjected to intensive structural studies (23)(24)(25), the crystal structure of the actin-Arp2/3 complex has eluded investigation so far. Before our study (26), the only available crystal structure of this kind was the yeast formin Bni1p FH2 domain that binds to two crystallographically related tetramethylrhodamine-modified actin (TMR-actin) subunits in a pseudo short-pitch fashion (27). However, the large size of TMR likely interferes with its interaction with actin and with actin Significance Actin filaments are the fundamental building blocks for cellular motility in muscle cells.…”

“…This strategy made possible the rapid determination of the first two crystal structures of oligomeric actin with tandem-Gactin-binding nucleator complexes: a mammalian nucleator Cordonbleu (Cobl) (26) and a bacterial effector Vibrio parahaemolyticus protein L (VopL) (28). Importantly, the observed non-filament-like conformation in actin-Cobl and the filament-like conformation in actin-VopL together suggest that both types of conformation are fully accessible to an actin complex obtained via the double-mutant strategy; thus the observed structure most likely reflects its native functional state.…”

Polypyrrole/TiO2 nanotube arrays with coaxial heterogeneous structure as sulfur hosts for lithium sulfur batteries

“…Fig. S9 is the discharge/charge capacity of 23 S/PPy/TiO2 NTs-300 cycled at various C-rates from 0.05 C to 1 C. After an initial discharge capacity of 1314.9 mAh g -1 at 0.05 C, the discharge capacity began to stabilize at about 1350 mAh g -1 . Further cycling at 0.1, 0.2, 0.5 and 1 C showed high reversible capacities of 1130, 950, 760 and 500 mAh g -1 , respectively.…”

“…Due to their high electrical conductivity along with sensibly designed framework to confine sulfur, trap soluble PS and eliminate volume expansion, these carbon materials have shown great improvements during the past decade. Besides, conductive polymers such as polypyrrole (PPy) [19][20][21], polyaniline (PANI) [22] and poly (3,4-(ethylenedioxy)thiophene) (PEDOT) [23] have also played a significant role in Li−S cells because of their elastic long chain structure, special functional groups and strong chemical bonds with sulfur. TiO2, as an inorganic material, is technically competent as a host for sulfur, but always was limited by its small surface area and pore volume until a sulfur-TiO2 yolk-shell nanoarchitecture proposed by Cui and coworkers [24].…”