  The hydroxyl-terminated self-assembled monolayer (OH-SAM), as a surface
resistant to protein adsorption, exhibits substantial potential in applications
such as ship navigation and medical implants, and the appropriate strategies
for designing anti-fouling surfaces are crucial. Here, we employ molecular
dynamics simulations and alchemical free energy calculations to systematically
analyze the factors influencing resistance to protein adsorption on the SAMs
terminated with single or double OH groups at three packing densities ({\Sigma}
= 2.0 nm-2, 4.5 nm-2, and 6.5 nm-2), respectively. For the first time, we
observe that the compactness and order of interfacial water enhance its
physical barrier effect, subsequently enhancing the resistance of SAM to
protein adsorption. Notably, the weak spatial hindrance effect of SAM leads to
the embedding of protein into SAM, resulting in a lack of resistance of SAM
towards protein. Furthermore, the number of hydroxyl groups per unit area of
double OH-terminated SAM at {\Sigma} = 6.5 nm-2 is approximately 2 to 3 times
that of single OH-terminated SAM at {\Sigma} = 6.5 nm-2 and 4.5 nm-2,
consequently yielding a weaker resistance of double OH-terminated SAM towards
protein. Meanwhile, due to the structure of SAM itself, i.e., the formation of
a nearly perfect ice-like hydrogen bond structure, the SAM exhibits the weakest
resistance towards protein. This study will complement and improve the
mechanism of OH-SAM resistance to protein adsorption, especially the
traditional barrier effect of interfacial water.
