Quantifying the effects of molecular orientation and length on two-dimensional receptor-ligand binding kinetics.
Surface presentation of adhesion receptors influences cell adhesion, although the mechanisms underlying these effects are not well understood. We used a micropipette adhesion frequency assay to quantify how the molecular orientation and length of adhesion receptors on the cell membrane affected two-dimensional kinetic rates of interactions with surface ligands. Interactions of P-selectin, E-selectin, and CD16A with their respective ligands or antibody were used to demonstrate such effects. Randomizing the orientation of the adhesion receptor or lowering its ligand- and antibody-binding domain above the cell membrane lowered two-dimensional affinities of the molecular interactions by reducing the forward rates but not the reverse rates. In contrast, the soluble antibody bound with similar three-dimensional affinities to cell-bound P-selectin constructs regardless of their orientation and length. These results demonstrate that the orientation and length of an adhesion receptor influences its rate of encountering and binding a surface ligand but does not subsequently affect the stability of binding.
National Microgravity Laboratory, Institute of Mechanics, Chinese Academy of Sciences, Beijing 100080, China.
Jun
J
Huang
Juan
J
Chen
Scott E
SE
Chesla
Tadayuki
T
Yago
Padmaja
P
Mehta
Rodger P
RP
McEver
Cheng
C
Zhu
Mian
M
Long
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D016466:CHO Cells; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D019040:E-Selectin / Q000737:chemistry; D004912:Erythrocytes / Q000378:metabolism; D018922:HL-60 Cells; D006801:Humans; D007700:Kinetics; D008024:Ligands; D008954:Models, Biological; D008956:Models, Chemical; D019007:P-Selectin / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017452:Receptors, IgG / Q000737:chemistry* / Q000378:metabolism; D013997:Time Factors
D000911:Antibodies, Monoclonal; D019040:E-Selectin; D008024:Ligands; D019007:P-Selectin; D017452:Receptors, IgG