On just a few nights every year, mother estuarine crabs rush down to the beach from their shore-side homes, ready to launch their young on their way. After carrying their precious cargo on tiny abdominal hairs and releasing them into the water, only the abandoned egg cases are left clinging to the mothers'ovigerious hairs. Soon after, the tightly attached cases loosen their grip,leaving the hairs free for the next clutch of eggs. But how the firmly attached cases are shed was a complete mystery, until Masayuki Saigusa began testing the water as the mothers cast their young out into the world. The water seemed to contain a protein that had the power to release the redundant egg cases from the female's body. But what was this mysterious polypeptide?Saigusa and his team set about isolating and cloning the protein to reveal its identity (p. 621).
Knowing that they could only collect the protein from egg-laden females,the team headed off to a roadside beach ready to trap the sea-bound mothers. Oleg Gusev remembers that the crabs were not particularly happy when intercepted, nipping at their captor before he could collect the case-detaching essence.
Back in the lab, the team separated the solutes from the water, testing each fraction on egg laden crab hairs to see which sample would loosen the bound cases. Sure enough, Saigusa found a sample from the water that dislodged the cases, and when he separated the protein components on an SDS-polyacrilamide gel, he discovered two protein chains. Sequencing short sections of the smaller 25 kDa peptide, Saigusa's team was able to get enough information on the short protein to clone it, and discover the protein's identity. It was a serine protease, making it a member of a well-known family of enzymes involved in processing other proteins.
But the team was in for a surprise. The serine protease gene that Gusev cloned seemed to code for a much larger protein than the 25 kDa serine protease that the team had isolated from the water. According to the gene, the egg detaching protein should weigh 54 kDa. What was going on? Why make a large protein when just a fragment seemed able to strip the abandoned cases from the hairs.
Gusev suspects that although the full-length protein contains the serine protease, the protease is only activated after the protein is cut in two. Gusev adds that he is also intrigued by the protein's remarkable environmental tolerance. He explains that most serine proteases are finely tuned, and only function under tightly regulated physiological conditions. But the crab's egg case-dislodging serine protease functions perfectly well, even when it's released into salty estuarine water.
The team is also curious to find which of the crab's tissues produce the protease. Gusev explains that both the mother and her young seem to produce the full length protein well before the youngsters hatch, so it's not clear whether the final dose of case clearing protease is delivered by the mother or her progeny. Either way, Saigusa and his team have their work cut out clearing up the case of the crab's self-cleaning protease.