X-ray Crystal Structure of the UCS Domain-Containing UNC-45 Myosin Chaperone from Drosophila melanogaster

Abstract

SUMMARY UCS proteins, such as UNC-45, influence muscle contraction and other myosin-dependent motile processes. We report the first x-ray crystal structure of a UCS domain-containing protein, the UNC-45 myosin chaperone from Drosophila melanogaster (DmUNC-45). The structure reveals that the Central and UCS domains form a contiguous arrangement of seventeen consecutive helical layers that arrange themselves into five discrete armadillo repeat subdomains. Small-angle x-ray scattering data suggest that free DmUNC-45 adopts an elongated conformation and exhibits flexibility in solution. Protease sensitivity maps to a conserved loop that contacts the most carboxy-terminal UNC-45 armadillo repeat sub-domain. Amino acid conservation across diverse UCS proteins maps to one face of this carboxy-terminal sub-domain and the majority of mutations that affect myosin-dependent cellular activities lie within or around this region. Our crystallographic, biophysical, and biochemical analyses suggest that DmUNC-45 function is afforded by its flexibility and by structural integrity of its UCS domain.

The myosin chaperone UNC45B is involved in lens development and autosomal dominant juvenile cataract

“…The majority of the mutations, including the human p.Arg805Trp mutation, are located in the UCS domain (Figure 1d). Lee et al 33 proposed a 3D structure from X-ray analysis of Drosophila melanogaster UNC-45, which we used to make a 3D model of the human protein (Figure 1d). 34 The structure reveals a contiguous arrangement of 17 helical layers with five discrete armadillo (ARM) repeat subdomains, which is folded to an L shape with no obvious transition between the central region and the UCS domain.…”

“…34 The structure reveals a contiguous arrangement of 17 helical layers with five discrete armadillo (ARM) repeat subdomains, which is folded to an L shape with no obvious transition between the central region and the UCS domain. 33 Five of the six known mutations in the UCS domain are located in a small nonpolar subregion, which has the highest degree of sequence identity in the five most evolutionarily divergent UCS proteins. This B40 amino-acid subregion forms the a-helices 18H1-3 and 19H1-2 ( Figure 1e) and with the exception of the steif nonsense allele, all the mutations in this subdomain are missense mutations.…”

“…In addition, the Arg803 position in Drosophila, which corresponds to h-Arg805 and C. elegans-Arg819, forms an inter-repeat ion pair interaction with Drosophila Glu766, which corresponds to h-Glu767 and C. elegansGlu781. 33 The C. elegans-Glu781 position is one of the ts paralyzing mutations, and the ion-interacting Arg residue is the C00116 Arg805Trp mutation. Both mutations may lead to ion pair instability of the a-helix structures (18H2 and 19H2 in Figure 1e) and reduced myosin-binding ability of the UCS domain.…”

“…Two hundred and fifty pg of 33 The five ARMR structures are shown in different colors with the conserved groove in the brown. The h-Glu768 and the h-Arg805 residues are shown by arrows.…”

“…Phyre2 and FirstGlance 34 were used for making the human model. (e) Alignment of the highly conserved subregion of the UCS domain with part of the a-helix structure for 18H-19H (modified from 33 ). Altered amino-acid positions are colored red and the conserved inter-repeat interacting ion pair h-Glu768 and h-Arg805 residues (Glu766 and Arg803 in the D. melanogaster sequence) are denoted.…”