Phospholipid determinants for annexin V binding sites and the role of tryptophan 187.
Annexin V is part of a family of Ca(2+)-dependent phospholipid-binding proteins, whose purported functions are related to their interactions with biological membranes. While Ca(2+)-dependent binding to phospholipids is well-established, the specific structural interactions within the phospholipid-binding sites have only been inferred to resemble those of phospholipase A2, with no direct structural evidence. In this study, the binding avidity of various phospholipid analogs, with variations at the headgroup or sn-2 acyl chain, was monitored in a C12E8 detergent micelle system using the increase in fluorescence of tryptophan 187. Micelles also contained excess negative surface charge to saturate a nonspecific component of the binding. The Ca2+ and phospholipid concentrations required for the binding of annexin V to various phospholipid headgroups were very similar, except for the relatively weak binding to phosphatidylinositol (PI). The unique close proximity of the PI sugar ring to the phosphate group may lead to steric hindrance in this case. Binding was also strongly dependent on the presence of an sn-3 phosphate group and an sn-2 acyl chain, as previously observed. The relatively shallow nature of the annexin V phospholipid-binding sites was reflected by the nearly equivalent binding of D and L versions of phospholipids, i.e., a large shift in the position of the sn-1 acyl chain is accommodated in this case. Binding of annexin V does not specifically require an ester carbonyl oxygen, as it occurs with ether-linked, amide-linked, and phosphonate-linked sn-2 hydrocarbon chains, under these conditions.(ABSTRACT TRUNCATED AT 250 WORDS)
Meers
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P
Department of Pathology, Boston University School of Medicinem Massachusetts 02118-2394.
Mealy
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T
D000215:Acylation; D000818:Animals; D017304:Annexin A5 / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D002417:Cattle; D008823:Micelles; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D013050:Spectrometry, Fluorescence; D013237:Stereoisomerism; D013329:Structure-Activity Relationship; D014364:Tryptophan / Q000378:metabolism*
D017304:Annexin A5; D008823:Micelles; D010743:Phospholipids; D014364:Tryptophan