Roles of individual domains of annexin I in its vesicle binding and vesicle aggregation: a comprehensive mutagenesis study.
To understand the mechanism by which annexin I induces membrane aggregation, a comprehensive mutagenesis of all six Ca2+-binding sites was performed. When the cap residues of type II Ca2+-binding sites were systematically mutated to Ala, a type II site in domain II was shown to be essential for Ca2+-dependent vesicle binding of annexin I. Domain II was not, however, directly involved in vesicle aggregation. Instead, type II sites in domains III and IV, respectively, and type III sites in domains I and IV were involved in vesicle aggregation. When all type II sites were deactivated, three type III sites provided residual vesicle binding and aggregating activities. Their contributions to these activities in the presence of type II sites were, however, relatively insignificant. To further investigate the role of each domain harboring a type II site, a set of mutants containing only a specific type II site(s) were generated and their activities measured. These measurements again underscored the importance of domain II in vesicle binding of annexin I and the involvement of domains III and IV in vesicle aggregation. The roles of individual domains in vesicle binding and aggregation can be accounted for by the conformational change of membrane-bound annexin I involving modular rotation of domains (I/IV) following the initial membrane adsorption of domains (II/III). In conjunction with mutagenesis studies on other annexins, these results show that individual domains of annexins, although structurally homologous, have distinct functions and that different annexins might interact with membranes via different domains.
Bitto
E
E
Department of Chemistry (M/C 111), University of Illinois at Chicago 60607-7061, USA.
Cho
W
W
D019943:Amino Acid Substitution / Q000235:genetics; D017305:Annexin A1 / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D002118:Calcium / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D008081:Liposomes / Q000737:chemistry* / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000138:chemical synthesis / Q000378:metabolism
D017305:Annexin A1; D008081:Liposomes; D011994:Recombinant Proteins; D002118:Calcium