Deletion of the N- or C-Terminal Helix of Apolipophorin III To Create a Four-Helix Bundle Protein

Biochemistry
Pankaj DwivediP M Weers

Abstract

Apolipophorin III (apoLp-III) is an exchangeable apolipoprotein found in insects and plays an important function in lipid transport. The protein has an unusual five-helix bundle architecture, deviating from the common four-helix bundle motif. To understand the role of the additional helix in apoLp-III, the N-terminal or C-terminal helix was deleted to create a putative four-helix bundle protein. While the protein lacking helix-1 could be expressed in bacteria albeit at reduced yields, apoLp-III lacking helix-5 could not be produced. Mutational analysis by truncating helix-5 showed that a minimum segment of approximately one-third of the C-terminal helix is required for protein expression. The variant lacking helix-5 was produced by inserting a methionine residue between helix-4 and -5; subsequent cyanogenbromide cleavage generated the four-helix variant. Both N- and C-terminal helix deletion variants displayed significantly reduced helical content, protein stability, and tertiary structure. Despite the significantly altered structure, the variants were still fully functional. The rate of dimyristoylphosphatidylcholine vesicle solubilization was enhanced 4-5-fold compared to the wild-type protein, and the deletion variants were ...Continue Reading

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Related Concepts

Apolipophorin-II
Metazoa
Apolipoproteins
LDL-1
Plasma Protein Binding Capacity
Protein Conformation
Mutagenesis, Site-Directed
Deletion Mutation
Locusta migratoria
Dimyristoylphosphatidylcholine

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