This article presents new insights into the molecular mechanism for regulating red cell membrane protein interactions that are responsible for erythrocyte membrane mechanical properties. For various skeletal proteins, structure-function correlations of protein 4.1R have been studied in detail. Kinetic analysis with the resonant mirror detection method has determined the nature of 4.1R interactions with various binding partners such as band 3, glycophorin C, and p55, and their binding sites. More importantly, calmodulin (CaM) binds to 4.1R in a Ca2+-independent manner to modulate the 4.1R interactions in the presence of Ca2+ at microM. Crystal structure of the 30-kD domain of 4.1R has a cloverleaf-like architecture with three lobes, each of which contains a binding region specific for binding partners. CaM binds to the grooves situated in two regions between the three lobes, possibly leading to conformational changes of the three lobes with a consequent alteration in the capacity of 4.1R to bind to its partners. The present findings on erythrocyte 4.1R should provide a basis for better understanding the membrane functions of nonerythroid cells.