An N-terminal three-helix fragment of the exchangeable insect apolipoprotein apolipophorin III conserves the lipid binding properties of wild-type protein.
Apolipophorin III (apoLp-III) from the greater wax moth Galleria mellonella is an exchangeable insect apolipoprotein that consists of five amphipathic alpha-helices, sharing high sequence identity with apoLp-III from the sphinx moth Manduca sexta whose structure is available. To define the minimal requirement for apoLp-III structural stability and function, a C-terminal truncated apoLp-III encompassing residues 1-91 of this 163 amino acid protein was designed. Far-UV circular dichroism spectroscopy revealed apoLp-III(1-91) has 50% alpha-helix secondary structure content in buffer (wild-type apoLp-III 86%), increasing to essentially 100% upon interactions with dimyristoylphosphatidylcholine (DMPC). Guanidine hydrochloride denaturation studies revealed similar stability properties for wild-type apoLp-III and apoLp-III(1-91). Resistance to denaturation for both proteins increased substantially upon association with phospholipid. In the absence of lipid, wild-type apoLp-III was monomeric whereas apoLp-III(1-91) partly formed dimers and trimers. Discoidal apoLp-III(1-91)-DMPC complexes were smaller in diameter (13.5 nm) compared to wild-type apoLp-III (17.7 nm), and more molecules of apoLp-III(1-91) associated with the complexes. Lipid interaction revealed that apoLp-III(1-91) binds to modified spherical lipoprotein surfaces and efficiently transforms phospholipid vesicles into discoidal complexes. Thus, the first three helices of G. mellonella apoLp-III contain the basic features required for maintenance of the structural integrity of the entire protein.
Institute of Biology/Zoology, Free University of Berlin, Königin-Luise-Strasse 1-3, D-14 195 Berlin, Germany.
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Dettloff
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Weers
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Niere
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Kay
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Ryan
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Wiesner
D000818:Animals; D001053:Apolipoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D017124:Conserved Sequence; D003432:Cross-Linking Reagents / Q000737:chemistry; D019476:Insect Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry; D008077:Lipoproteins, LDL / Q000378:metabolism; D046911:Macromolecular Substances; D009036:Moths; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000138:chemical synthesis / Q000302:isolation & purification / Q000648:ultrastructure; D017384:Sequence Deletion; D013499:Surface Properties; D010738:Type C Phospholipases / Q000737:chemistry
D001053:Apolipoproteins; D003432:Cross-Linking Reagents; D019476:Insect Proteins; D008055:Lipids; D008077:Lipoproteins, LDL; D046911:Macromolecular Substances; D010446:Peptide Fragments; D010743:Phospholipids; D011994:Recombinant Proteins; C042470:apolipophorin III; D010738:Type C Phospholipases