TY - JOUR

T1 - Hansenula polymorpha Pex3p is a peripheral component of the peroxisomal membrane

AU - Haan, Gert Jan

AU - Faber, Klaas Nico

AU - Baerends, Richard J S

AU - Koek, Anne

AU - Krikken, Arjen

AU - Kiel, Jan A K W

AU - van der Klei, Ida J

AU - Veenhuis, Marten

N1 - Relation: https://www.rug.nl/gbb/ date_submitted:2007 Rights: University of Groningen, Groningen Biomolecular Sciences and Biotechnology Institute

PY - 2002/7/19

Y1 - 2002/7/19

N2 - Hansenula polymorpha Pex3p plays an essential role in the biogenesis and maintenance of the peroxisomal membrane. In the initial report, bakers' yeast Pex3p was suggested to represent an integral component of the peroxisomal membrane, containing one membrane-spanning region that exposes the N terminus of the protein into the organellar matrix. Biochemically, HpPex3p behaved like an integral membrane protein as it was resistant toward high salt and carbonate treatment. However, urea fully removed Pex3p from the membrane under conditions in which the integral membrane protein Pex10p was resistant to this treatment. Additional experiments, including protease protection assays and pre-embedding labeling experiments on purified organellar fractions from cells that produced Pex3ps carrying Myc epitopes at various selected locations in the protein, revealed that invariably all Myc tags were accessible for externally added proteases and antibodies, independent of the presence of detergents. Also, overproduction of Pex3p failed to demonstrate the typical integral membrane protein structures in fracture faces of freeze-fractured peroxisomes. Taken together, our data suggest that HpPex3p does not span the peroxisomal membrane but instead is tightly associated to the cytosolic face of the organelle where it may be present in focal protein clusters.

AB - Hansenula polymorpha Pex3p plays an essential role in the biogenesis and maintenance of the peroxisomal membrane. In the initial report, bakers' yeast Pex3p was suggested to represent an integral component of the peroxisomal membrane, containing one membrane-spanning region that exposes the N terminus of the protein into the organellar matrix. Biochemically, HpPex3p behaved like an integral membrane protein as it was resistant toward high salt and carbonate treatment. However, urea fully removed Pex3p from the membrane under conditions in which the integral membrane protein Pex10p was resistant to this treatment. Additional experiments, including protease protection assays and pre-embedding labeling experiments on purified organellar fractions from cells that produced Pex3ps carrying Myc epitopes at various selected locations in the protein, revealed that invariably all Myc tags were accessible for externally added proteases and antibodies, independent of the presence of detergents. Also, overproduction of Pex3p failed to demonstrate the typical integral membrane protein structures in fracture faces of freeze-fractured peroxisomes. Taken together, our data suggest that HpPex3p does not span the peroxisomal membrane but instead is tightly associated to the cytosolic face of the organelle where it may be present in focal protein clusters.

KW - ATP-Binding Cassette Transporters

KW - Base Sequence

KW - Fungal Proteins

KW - Intracellular Membranes

KW - Membrane Proteins

KW - Microscopy, Electron

KW - Microscopy, Fluorescence

KW - Molecular Sequence Data

KW - Peroxisomes

KW - Pichia

KW - Saccharomyces cerevisiae Proteins

U2 - 10.1074/jbc.M108569200

DO - 10.1074/jbc.M108569200

M3 - Article

C2 - 12011037

VL - 277

SP - 26609

EP - 26617

JO - The Journal of Biological Chemistry

JF - The Journal of Biological Chemistry

SN - 0021-9258

IS - 29

ER -