Phosphate-Driven\nInterfacial Self-Assembly of Silk\nFibroin for Continuous Noncovalent Growth of Nanothin Defect-Free\nCoatings

Silk\nfibroin is a fiber-forming protein derived from the thread\nof <i>Bombyx mori</i> silkworm cocoons. This biocompatible\nprotein, under the kosmotropic influence of potassium phosphate, can\nundergo supramolecular self-assembly driven by a random coil to β-sheet\nsecondary structure transition. By leveraging concurrent nonspecific\nadsorption and self-assembly of silk fibroin, we demonstrate an interfacial\nphenomenon that yields adherent, defect-free nanothin protein coatings\nthat grow continuously in time, without observable saturation in mass\ndeposition. This noncovalent growth of silk fibroin coatings is a\ndeparture from traditionally studied protein adsorption phenomena,\nwhich generally yield adsorbed layers that saturate in mass with time\nand often do not completely cover the surface. Here, we explore the\nfundamental mechanisms of coating growth by examining the effects\nof coating solution parameters that promote or inhibit silk fibroin\nself-assembly. Results show a strong dependence of coating kinetics\nand structure on solution pH, salt species, and salt concentration.\nMoreover, coating growth was observed to occur in two stages: an early\nstage driven by protein–surface interactions and a late stage\ndriven by protein–protein interactions. To describe this phenomenon,\nwe developed a kinetic adsorption model with Langmuir-like behavior\nat early times and a constant steady-state growth rate at later times.\nStructural analysis by FTIR and photoinduced force microscopy show\nthat small β-sheet-rich structures serve as anchoring sites\nfor absorbing protein nanoaggregates, which is critical for coating\nformation. Additionally, β-sheets are preferentially located\nat the interface between protein nanoaggregates in the coating, suggesting\ntheir role in forming stable, robust coatings.