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J. Biol. Chem., Vol. 280, Issue 52, 42580-42591, December 30, 2005

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Calcium Triggers Folding of Lipoprotein Lipase into Active Dimers^*

Liyan Zhang, Aivar Lookene, Gengshu Wu1, and Gunilla Olivecrona2

From the Department of Medical Biosciences, Physiological Chemistry, Umeå University, SE-901 87 Umeå, Sweden and the Department of Gene Technology, Tallinn Technical University, Tallinn 12618, Estonia

The active form of lipoprotein lipase (LPL) is a noncovalent homodimer of 55-kDa subunits. The dimer is unstable and tends to undergo irreversible dissociation into inactive monomers. We noted that a preparation of such monomers slowly regained traces of activity under assay conditions with substrate, heparin, and serum or in cell culture medium containing serum. We therefore studied the refolding pathway of LPL after full denaturation in 6 M guanidinium chloride or after dissociation into monomers in 1 M guanidinium chloride. In crude systems, we identified serum as the factor promoting reactivation. Further investigations demonstrated that Ca²⁺ was the crucial component in serum for reactivation of LPL and that refolding involved at least two steps. Studies of far-UV circular dichroism, fluorescence, and proteolytic cleavage patterns showed that LPL started to refold from the C-terminal domain, independent of calcium. The first step was rapid and resulted in formation of an inactive monomer with a completely folded C-terminal domain, whereas the N-terminal domain was in the molten globule state. The second step was promoted by Ca²⁺ and converted LPL monomers from the molten globule state to dimerization-competent and more tightly folded monomers that rapidly formed active LPL dimers. The second step was slow, and it appears that proline isomerization (rather than dimerization as such) is rate-limiting. Inactive monomers isolated from human tissue recovered activity under the influence of Ca²⁺. We speculate that Ca²⁺-dependent control of LPL dimerization might be involved in the normal post-translational regulation of LPL activity.

Received for publication, July 5, 2005 , and in revised form, September 6, 2005.

^* This work was supported by Swedish Research Council Grant 12203 and by a Swedish Royal Academy of Sciences scholarship (to A. L.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ Present address: Dept. of Biochemistry, University of Alberta, Edmonton, Alberta T6G 2S2, Canada.

² To whom correspondence should be addressed: Dept. of Medical Biosciences, Physiological Chemistry, Umeå University, Bldg. 6M, 3rd Floor, SE-901 87 Umeå, Sweden. Tel.: 46-90-785-4491; Fax: 46-90-785-4496; E-mail: Gunilla.Olivecrona{at}medbio.umu.se.

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