Oligo(ethylene glycol) (O-EG(n))-terminated alkanethiol surface-assembled monolayers (SAMs) have been reported to demonstrate protein-resistant properties similar to those of poly(ethylene glycol) (PEG). In this study, we compared the relative protein resistance of short and long ethylene oxide chains, SAMs of PEG 5000, PEG 2000, O-EG(3) (molecular weight = 120), and O-EG(6) (molecular weight = 240), on gold surfaces. Surface plasmon resonance showed that these monolayers were all protein-resistant within the uncertainty of the measurement. However, they exhibited different adhesive properties toward 3T3 mouse fibroblast adhesion in supplemented Dulbecco's modified Eagles medium. The results show that the cell adhesion was sensitive to the concentration of proteins supplemented in the culture medium and to the length of PEG chains.

The structures of many cell surface adhesion proteins comprise multiple tandem repeats of structurally similar domains. In many cases, the functional significance of this architecture is unknown, and there are several cases in which evidence for individual domain involvement in adhesion has been contradictory. In particular, the extracellular region of the adhesion glycoprotein cadherin consists of five tandemly arranged domains. One proposed mechanism postulated that adhesion involves only trans interactions between the outermost domains. However, subsequent investigations have generated several competing models. Here we describe direct measurements of the distance-dependent interaction potentials between cadherin mutants lacking different domains. By quantifying both the absolute distances at which opposed cadherin fragments bind and the quantized changes in the interaction potentials that result from deletions of individual domains, we demonstrate that two domains participate in homophilic cadherin binding. This finding contrasts with the current view that cadherins bind via a single, unique site on the protein surface. The potentials that result from interactions involving multiple domains generate a novel, modular binding mechanism in which opposed cadherin ectodomains can adhere in any of three antiparallel alignments.

Direct force measurements were used to investigate the molecular mechanism of heterophilic adhesion between the murine T-cell adhesion glycoprotein CD2 and its ligand CD48. From the distance dependence of the protein-protein interaction potential, we demonstrate directly that the full-length extracellular domains adhere in a head-to-head orientation. The absence of long-range electrostatic protein-protein attraction further indicates that the salt bridges between the binding surfaces only influence the interaction at short range. Despite the loss of a stabilizing disulfide bond in domain 1 (D1) of CD2, adhesive failure occurs abruptly with no evidence of partial protein unfolding during detachment. Finally, these measurements between extended membrane surfaces directly confirm that the low-affinity CD2-CD48 bond generates weak adhesion and that lateral receptor mobility is required for the development of appreciable adhesion. This is the first direct measurement of the range and magnitude of the forces governing heterotypic adhesion mediated by cell surface proteins. These results both verified the head-to-head CD2-CD48 docking alignment and demonstrated the ability to elucidate the structure-function relationships of adhesion proteins from the measured distance dependence of their interaction potentials.

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