Abstract
Phosphatidylserine (PS) on apoptotic cells is a target for diagnosis and therapy using annexin A5 (anxA5). Pretargeting is a strategy developed to improve signal to background ratio for molecular imaging and to minimize undesired side effects of pharmacological and radiotherapy. Pretargeting relies on accessibility of the target finder on the surface of the target cell. anxA5 binds PS and crystallizes in a two-dimensional network covering the PS-expressing cell surface. Two-dimensional crystallization is the driving force for anxA5 internalization by PS-expressing cells. Here, we report structure/function analysis of anxA5 internalization. Guided by structural bioinformatics including protein-protein docking, we revealed that the amino acids Arg(63), Lys(70), Lys(101), Glu(138), Asp(139), and Asn(160) engage in intermolecular salt bridges within the anxA5 trimer, which is the basic building block of the two-dimensional network. Disruption of the salt bridges by site-directed mutagenesis does not affect PS binding but inhibits trimer formation and cell entry of surface-bound anxA5. The anxA5 variants with impaired internalization are superior molecular imaging agents in pretargeting strategies as compared with wild-type anxA5.
Highlights
Cell surface-expressed phosphatidylserine (PS)2 is an attractive biomarker for diagnosis of diseases and evaluation of efficacy of treatment [1]
We embarked on a study to unravel the relationship between annexin A5 (anxA5) structure and two-dimensional crystallization to generate anxA5 variants that bind cell surface expressed PS without opening the portal of cell entry
We have investigated the structure/function relationship of anxA5 internalization by cells executing apoptosis
Summary
Cell surface-expressed phosphatidylserine (PS)2 is an attractive biomarker for diagnosis of diseases and evaluation of efficacy of treatment [1]. AnxA5 binds PS and crystallizes in a two-dimensional network covering the PS-expressing cell surface. AnxA5 binds cell surface expressed PS and induces membrane invagination through a nanomechanical mechanism involving the twodimensional crystallization of anxA5 trimers [13].
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