Crystal structure of the BstDEAD N-terminal domain: A novel DEAD protein from Bacillus stearothermophilus

Abstract

Most cellular processes requiring RNA structure rearrangement necessitate the action of Asp-Glu-Ala-Asp (DEAD) proteins. Members of the family, named originally for the conserved DEAD amino acid sequence, are thought to disrupt RNA structure and facilitate its rearrangement by unwinding short stretches of duplex RNA. BstDEAD is a novel 436 amino acid representative of the DEAD protein family from Bacillus stearothermophilus that contains all eight conserved motifs found in DEAD proteins and is homologous with other members of the family. Here, we describe the 1.85 Å resolution structure of the N-terminal domain (residues 1-211) of BstDEAD (BstDEAD-NT). Similar to the corresponding domains of related helicases, BstDEAD-NT adopts a parallel ␣/␤ structure with RecA-like topology. In general, the conserved motifs superimpose on closely related DEAD proteins and on more distantly related helicases such as RecA. This affirms the current belief that the core helicase domains, responsible for mechanistic activity, are structurally similar in DEAD proteins. In contrast, however, the so-called Walker A P-loop, which binds the ␤-and ␥-phosphates of ATP, adopts a rarely seen "closed" conformation that would sterically block ATP binding. The closed conformation may be indicative of a general regulatory feature among DEAD proteins (and RNA helicases) that differs from that used by DNA helicases. BstDEAD also contains a unique extension of ∼ 60 residues at the C terminus that is highly basic, suggesting that it might bind nucleic acids and, in so doing, confer specificity to the helicase activity of the core region.

Crystal structure and functional analysis of DEAD-box protein Dhh1p

“…In tDhh1p structure, the P loop adopts an ''open'' conformation ( Fig. 3C) similar to that observed in mjDEAD, but opposed to the ''closed conformation'' in the structures of eIF4A and BstDEAD without bound ligand (Johnson and McKay 1999;Story et al 2001;Carmel and Matthews 2004). Such an open conformation of motif I would facilitate ATP binding.…”

“…Despite the large number of DEAD-box proteins from bacteria to human and their universal roles in so many processes, high-resolution structures remain limited to three DEAD proteins, yeast eIF4A, a prototype minimal DEAD-box protein (Benz et al 1999;Johnson and McKay 1999;Caruthers et al 2000); mjDEAD from Methanococcus janaschii (Story et al 2001); and the N-terminal domain of BstDEAD from Bacillus stearothermophilus (Carmel and Matthews 2004). Recently, the crystal structure of UAP56, a DExD/H-box protein involved in pre-mRNA splicing and mRNA export, revealed a unique spatial arrangement of the two conserved helicase domains (Shi et al 2004).…”

The DEAD-box protein family of RNA helicases

“…Notably, this includes the SF1 PcrA (Subramanya et al, 1996;Velankar et al, 1999), SF2 UvrB (Theis et al, 1999), SF1 Rep (Korolev et al, 1997), the helicase domain of SF2 NS3 from the Hepatitis C virus ( Fig. 2A; Yao et al, 1997;Kim et al, 1998) and four SF2 DEAD-box proteins (Benz et al, 1999;Caruthers et al, 2000;Story et al, 2001;Carmel and Matthews, 2004;Shi et al, 2004;Zhao et al, 2004). Among the DEAD-box proteins, the crystal structures of the functionally uncharacterized MjDEAD from Methanococcus jannashii ( Fig.…”

“…Finally, the amino-terminal domain of BstDEAD was also crystallized and solved (Fig. 3E;Carmel and Matthews, 2004). All these DEAD-box proteins lack long amino-and carboxylterminal extensions, and they are close to the minimal size constituting the helicase core alone.…”

“…These additional structures correspond to, and encompass, the recently discovered Q motif that is specific to the DEAD-box protein family (see Section 3.2). The Q motif and the other conserved motifs (I to VI) are typically located at β-strand-loop or helixloop transitions (Tanner and Linder, 2001;Carmel and Matthews, 2004), as is true in the other helicase families.…”