Am J Physiol Cell Physiol AJP: Lung Cellular and Molecular Physiology
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Am J Physiol Cell Physiol 287: C1623-C1635, 2004. First published August 11, 2004; doi:10.1152/ajpcell.00142.2004
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CELLULAR METABOLISM

The behavior of peroxisomes in vitro: mammalian peroxisomes are osmotically sensitive particles

Vasily D. Antonenkov,1 Raija T. Sormunen,2 and J. Kalervo Hiltunen1

1Department of Biochemistry and Biocenter Oulu and 2Department of Pathology, University of Oulu, FIN-90014 Oulu, Finland

Submitted 16 March 2004 ; accepted in final form 5 August 2004

It has been known for a long time that mammalian peroxisomes are extremely fragile in vitro. Changes in the morphological appearance and leakage of proteins from purified particles demonstrate that peroxisomes are damaged during isolation. However, some properties of purified peroxisomes, e.g., the latency of catalase, imply that their membranes are not disrupted. In the current study, we tried to ascertain the mechanism of this unusual behavior of peroxisomes in vitro. Biochemical and morphological examination of isolated peroxisomes subjected to sonication or to freezing and thawing showed that the membrane of the particles seals after disruption, restoring permeability properties. Transient damage of the membrane leads to the formation of peroxisomal "ghosts" containing nucleoid but nearly devoid of matrix proteins. The rate of leakage of matrix proteins from broken particles depended inversely on their molecular size. The effect of polyethylene glycols on peroxisomal integrity indicated that these particles are osmotically sensitive. Peroxisomes suffered an osmotic lysis during isolation that was resistant to commonly used low-molecular-mass osmoprotectors, e.g., sucrose. Damage to peroxisomes was partially prevented by applying more "bulky" osmoprotectors, e.g., polyethylene glycol 1500. A method was developed for the isolation of highly purified and nearly intact peroxisomes from rat liver by using polyethylene glycol 1500 as an osmoprotector.

osmolarity; cell fractionation; isolation of organelles


Address for reprint requests and other correspondence: V. D. Antonenkov or J. Kalervo Hiltunen, Dept. Biochemistry and Biocenter Oulu; Univ. of Oulu, PO Box 3000, FIN-90014 Oulu, Finland (E-mail: vasily.antonenkov{at}oulu.fi or kalervo.hiltunen{at}oulu.fi)




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