PDBsum entry 4w7s

Hydrolase

PDB id: 4w7s

Contents

Protein chains

455 a.a.

Ligands

P6G ×8

GOL

ANP

Metals

_MG ×2

Waters ×251

PDB id:

4w7s

Name:

Hydrolase

Title:

Crystal structure of the yeast dead-box splicing factor prp28 at 2.54 angstroms resolution

Structure:

Pre-mRNA-splicing atp-dependent RNA helicase prp28. Chain: a, b. Synonym: helicase ca8. Engineered: yes

Source:

Saccharomyces cerevisiae. Baker's yeast. Organism_taxid: 559292. Strain: atcc 204508 / s288c. Gene: prp28, ydr243c, yd8419.10c. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.54Å R-factor: 0.169 R-free: 0.219

Authors:

A.Jacewicz,P.Smith,B.Schwer,S.Shuman

Key ref:

A.Jacewicz et al. (2014). Crystal structure, mutational analysis and RNA-dependent ATPase activity of the yeast DEAD-box pre-mRNA splicing factor Prp28. Nucleic Acids Res, 42, 12885-12898. PubMed id: 25303995 DOI: 10.1093/nar/gku930

Date:

22-Aug-14 Release date: 29-Oct-14

Protein chains

P23394  (PRP28_YEAST) -  Pre-mRNA-splicing ATP-dependent RNA helicase PRP28 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 588 a.a.

Struc: 455 a.a.

Enzyme reactions

• Enzyme class: E.C.3.6.4.13 - Rna helicase.
• Reaction: ATP + H2O = ADP + phosphate + H⁺

ATP + H2O = ADP (Bound ligand (Het Group name = ANP) matches with 81.25% similarity) + phosphate + H(+)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1093/nar/gku930

Nucleic Acids Res 42:12885-12898 (2014)

PubMed id: 25303995

Crystal structure, mutational analysis and RNA-dependent ATPase activity of the yeast DEAD-box pre-mRNA splicing factor Prp28.

A.Jacewicz, B.Schwer, P.Smith, S.Shuman.

ABSTRACT

Yeast Prp28 is a DEAD-box pre-mRNA splicing factor implicated in displacing U1 snRNP from the 5' splice site. Here we report that the 588-aa Prp28 protein consists of a trypsin-sensitive 126-aa N-terminal segment (of which aa 1-89 are dispensable for Prp28 function in vivo) fused to a trypsin-resistant C-terminal catalytic domain. Purified recombinant Prp28 and Prp28-(127-588) have an intrinsic RNA-dependent ATPase activity, albeit with a low turnover number. The crystal structure of Prp28-(127-588) comprises two RecA-like domains splayed widely apart. AMPPNP•Mg(2+) is engaged by the proximal domain, with proper and specific contacts from Phe194 and Gln201 (Q motif) to the adenine nucleobase. The triphosphate moiety of AMPPNP•Mg(2+) is not poised for catalysis in the open domain conformation. Guided by the Prp28•AMPPNP structure, and that of the Drosophila Vasa•AMPPNP•Mg(2+)•RNA complex, we targeted 20 positions in Prp28 for alanine scanning. ATP-site components Asp341 and Glu342 (motif II) and Arg527 and Arg530 (motif VI) and RNA-site constituent Arg476 (motif Va) are essential for Prp28 activity in vivo. Synthetic lethality of double-alanine mutations highlighted functionally redundant contacts in the ATP-binding (Phe194-Gln201, Gln201-Asp502) and RNA-binding (Arg264-Arg320) sites. Overexpression of defective ATP-site mutants, but not defective RNA-site mutants, elicited severe dominant-negative growth defects.