HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Seedorf, U. Articles by Assmann, G. Search for Related Content PubMed PubMed Citation Articles by Seedorf, U. Articles by Assmann, G. Social Bookmarking                      What's this?

J. Biol. Chem., Vol. 269, Issue 4, 2613-2618, Jan, 1994

Structure-activity studies of human sterol carrier protein 2

U Seedorf, S Scheek, T Engel, C Steif, HJ Hinz and G Assmann
Institut fur Arterioskleroseforschung, Westfalische Wilhelms- Universitat, Munster, Germany.

Recombinant human sterol carrier protein 2 (SCP2) variants were generated by site-directed mutagenesis and expression in Escherichia coli. The ability of the variants to stimulate microsomal conversion of 7-dehydrocholesterol to cholesterol (sterol carrier activity) and to transfer cholesterol and phosphatidylcholine from donor small unilamellar vesicles to acceptor membranes (cholesterol and phosphatidylcholine transfer activities) was compared with wild-type recombinant SCP2. Our results indicate that all measured activities of recombinant human pre-SCP2 (including the 20-amino acid leader sequence) and mature SCP2 were similar. Expressed glutathione S- transferase fusion proteins (GST-SCP2 and GST-pre-SCP2) possessed considerable activity, suggesting that steric obstruction at the amino terminus causes only minor inactivation. The effect of progressive removal of peptides from the carboxyl terminus showed that amino acids between Lys100 and Asn104 are essential for SCP2 activity. This conclusion was substantiated by the observation that replacing Asn104 with Asp or Ile caused considerable inactivation, whereas replacing Met105 with Leu had almost no effect. Since N-ethylmaleimide is known to inhibit SCP2 activity, substitutions were also introduced in the vicinity of Cys71. Whereas Val71 and Ser71 variants possessed wild-type activity, replacing Asp70 with Asn almost completely abolished SCP2 activity. Further, the importance of residues located close to the amino terminus was indicated by complete inactivation of a 10-amino- terminal amino acid deletion mutant and by replacing Leu20 with Glu. Circular dichroism results showed that Leu20 and Asp70 may serve to stabilize the overall fold, whereas residue 104 appears to play a role in the specific lipid binding and/or transfer activity of SCP2.
CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				V. J. J. Martin and W. W. Mohn Genetic Investigation of the Catabolic Pathway for Degradation of Abietane Diterpenoids by Pseudomonas abietaniphila BKME-9 J. Bacteriol., July 1, 2000; 182(13): 3784 - 3793. [Abstract] [Full Text]

				E. J. Murphy, T. Stiles, and F. Schroeder Sterol carrier protein-2 expression alters phospholipid content and fatty acyl composition in L-cell fibroblasts J. Lipid Res., May 1, 2000; 41(5): 788 - 796. [Abstract] [Full Text]

				N. Stolowich, A. Frolov, A. D. Petrescu, A. I. Scott, J. T. Billheimer, and F. Schroeder Holo-sterol Carrier Protein-2. 13C NMR INVESTIGATION OF CHOLESTEROL AND FATTY ACID BINDING SITES J. Biol. Chem., December 10, 1999; 274(50): 35425 - 35433. [Abstract] [Full Text] [PDF]

				B. P. Atshaves, A. D. Petrescu, O. Starodub, J. B. Roths, A. B. Kier, and F. Schroeder Expression and intracellular processing of the 58 kDa sterol carrier protein-2/3-oxoacyl-CoA thiolase in transfected mouse L-cell fibroblasts J. Lipid Res., April 1, 1999; 40(4): 610 - 622. [Abstract] [Full Text]

				E. J. Murphy Sterol carrier protein-2 expression increases NBD-stearate uptake and cytoplasmic diffusion in L cells Am J Physiol Gastrointest Liver Physiol, August 1, 1998; 275(2): G237 - G243. [Abstract] [Full Text] [PDF]

				U. Seedorf, M. Raabe, P. Ellinghaus, F. Kannenberg, M. Fobker, T. Engel, S. Denis, F. Wouters, K. W.A. Wirtz, R. J.A. Wanders, et al. Defective peroxisomal catabolism of branched fatty acyl coenzyme A in mice lacking the sterol carrier protein-2/sterol carrier protein-x gene function Genes & Dev., April 15, 1998; 12(8): 1189 - 1201. [Abstract] [Full Text]

				F. Leenders, J. G. Tesdorpf, M. Markus, T. Engel, U. Seedorf, and J. Adamski Porcine 80-kDa Protein Reveals Intrinsic 17beta-Hydroxysteroid Dehydrogenase, Fatty Acyl-CoA-hydratase/Dehydrogenase, and Sterol Transfer Activities J. Biol. Chem., March 8, 1996; 271(10): 5438 - 5442. [Abstract] [Full Text] [PDF]