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PMID: 22753491 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't

Structural and functional characterization of the two phosphoinositide binding sites of PROPPINs, a β-propeller protein family.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 109 ·No. 30 ·2012-07-24 ·Pages E2042-9

Krick R, Busse RA, Scacioc A, Stephan M, Janshoff A, Thumm M, Kühnel K

Abstract

β-propellers that bind polyphosphoinositides (PROPPINs), a eukaryotic WD-40 motif-containing protein family, bind via their predicted β-propeller fold the polyphosphoinositides PtdIns3P and PtdIns(3,5)P(2) using a conserved FRRG motif. PROPPINs play a key role in macroautophagy in addition to other functions. We present the 3.0-Å crystal structure of Kluyveromyces lactis Hsv2, which shares significant sequence homologies with its three Saccharomyces cerevisiae homologs Atg18, Atg21, and Hsv2. It adopts a seven-bladed β-propeller fold with a rare nonvelcro propeller closure. Remarkably, in the crystal structure, the two arginines of the FRRG motif are part of two distinct basic pockets formed by a set of highly conserved residues. In comprehensive in vivo and in vitro studies of ScAtg18 and ScHsv2, we define within the two pockets a set of conserved residues essential for normal membrane association, phosphoinositide binding, and biological activities. Our experiments show that PROPPINs contain two individual phosphoinositide binding sites. Based on docking studies, we propose a model for phosphoinositide binding of PROPPINs.

MeSH Terms
Amino Acid Motifs/genetics Autophagy-Related Proteins Binding Sites/genetics Cloning, Molecular Conserved Sequence/genetics Crystallography, X-Ray Kluyveromyces/chemistry Membrane Proteins/chemistry,genetics Models, Molecular Molecular Dynamics Simulation Mutagenesis Phosphatidylinositols/metabolism Protein Binding Protein Conformation Saccharomyces cerevisiae Proteins/chemistry,genetics
Chemicals
ATG18 protein, S cerevisiae Autophagy-Related Proteins Membrane Proteins Phosphatidylinositols Saccharomyces cerevisiae Proteins
Authors & Affiliations
7 authors, click to expand affiliations / ORCID
Krick Roswitha
Department of Biochemistry II, Georg August University, D-37073 Göttingen, Germany.
Busse Ricarda A
Scacioc Andreea
Stephan Milena
Janshoff Andreas
Thumm Michael
Kühnel Karin
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
1091-6490
DOI
10.1073/pnas.1205128109
Published
2012-07-24
Epub
2012-00-02
Pages
E2042-9
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC3409749
Subset
IM
Databases
PDB
4AV8, 4AV9
External Links
PubMed source DOI: 10.1073/pnas.1205128109 PMC Full Text View journal details
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