Carbonic anhydrase generates CO2 and H+ that drive spider silk formation via opposite effects on the terminal domains.

Abstract

Author SummaryThe spinning process of spider silk is crucial for making webs or other complex constructions to catch spider's prey. The main components of the silk are spidroins, which are large and repetitive proteins that have conserved nonrepetitive terminal domains (NT and CT). Spiders manage both to store the highly aggregation-prone spidroins in solution at extreme concentrations in the silk glands and then to rapidly convert these spidroins into a solid fiber within fractions of a second as they spin fibres. This process has been extensively studied and is thought to involve a pH gradient, but how this pH gradient is generated and maintained was not resolved. Here, we measured the pH at locations along the ampullate gland and determined that the pH decreases to 5.7 in the middle of the spinning duct. We also observed that the carbon dioxide pressure is simultaneously increased and that its accumulation may affect the stability of CT. We find that active carbonic anhydrase (CA) is crucial to maintain the pH gradient along the gland. Detailed molecular studies of NT and CT under the disparate conditions present along the gland revealed a lock and trigger mechanism whereby in more neutral pH conditions, precocious spidroin aggregation is prevented, and when in more acidic pH conditions, NT dimers firmly interconnect the spidroins and the CT unfolds into β-sheet nuclei that can trigger rapid polymerization of the spidroins. We conclude that this mechanism enables temporal and spatial control of silk formation and may be harnessed in attempts to produce artificial silk replicas.