HARDER, W

SULTER, GJ

VANDERKLEI, IJ

VEENHUIS, M

We have studied the expression of alcohol oxidase (AO) in a peroxisome-deficient mutant strain of Hansenula polymorpha. High levels of octameric, active AO (up to 3.0 U/mg protein) were detected in cells grown at low dilution rates in a glucose-limited chemostat in the presence of choline as the sole nitrogen source. Monomeric or other intermediate forms of AO were not detected in the mutant strain. This indicated that assembly of the protein into active octameric molecules in the cytosol was as efficient as in wild-type cells where this process is confined to the peroxisomal matrix. At relatively low rates of expression (less than 1 U/mg protein) AO was localized throughout the cytosol and, surprisingly, was also present inside the nucleus. However, at enhanced levels large crystalloids were formed. Generally one crystalloid was observed per cell, whereas smaller ones were occasionally found in developing buds. Also large crystalloids have been observed inside the nucleus. These crystalloids were not surrounded by a membrane. Based on the morphology of the molecules that constituted these crystalloids and the results of (immuno)cytochemical experiments we conclude that the crystalloids are composed of octameric AO molecules, arranged in a regular lattice, identical to the 3-dimensional architecture previously described for the crystalline matrix of peroxisomes in methanol-grown wild type cells of H. polymorpha. Attempts to purify the crystalloids by conventional fractionation methods failed, due to their apparent fragility; however, (immuno)cytochemical experiments revealed that catalase and dihydroxyacetone synthase were also associated with these structures.</p

Dissertations of the University of Groningen

ASSEMBLY OF ALCOHOL OXIDASE IN THE CYTOSOL OF A PEROXISOME-DEFICIENT MUTANT OF HANSENULA-POLYMORPHA - PROPERTIES OF THE PROTEIN AND ARCHITECTURE OF THE CRYSTALS

ASSEMBLY OF ALCOHOL OXIDASE IN THE CYTOSOL OF A PEROXISOME-DEFICIENT MUTANT OF HANSENULA-POLYMORPHA - PROPERTIES OF THE PROTEIN AND ARCHITECTURE OF THE CRYSTALS

Abstract

Similar works

Full text

Available Versions

Dissertations of the University of Groningen