The CEACAM1 N-terminal Ig domain mediates cis- and trans-binding and is essential for allosteric rearrangements of CEACAM1 microclusters

Klaile, Esther; Vorontsova, Olga; Sigmundsson, Kristmundur; Müller, Mario M.; Singer, Bernhard B.; Öfverstedt, Lars-Göran; Svensson, Stina; Skoglund, Ulf; Öbrink, Björn

Individual Ig domains were resolved in many of the tomographically determined CEACAM1 ectodomains ( Fig. 4 and Video 1). Because the rat CEACAM1 D1 domain is larger and has more glycosylation sites (105 amino acids and three N-glycosylation sites) than the D4 domain (67 amino acids and two N-glycosylation sites; Edlund et al., 1993) , it might be possible to distinguish the two ends in individual molecules. To test this possibility, 20 different, extended monomeric D(1-4) domains were divided into four equally long segments, and the mean of the two diameters of the ellipsoidal center cross sections of each segment was recorded. This showed that one end had a significantly larger mean diameter than the other end in all molecules. Setting the mean of the larger end mean diameters to 1 gave a mean of the smaller end mean diameters of 0.81 ± 0.09 (P = 4 × 10 11 ). Similar analysis of D(2-4), which was divided into three segments, showed no significant difference in the size of the two end domains, the smaller having a relative size of 0.96 ± 0.05 (P = 0.09; D2 has 74 amino acids and six N-glycosylation sites). This indicates that the D1 and D4 ends can be discriminated in a large proportion of the recorded molecules.