A gene that encodes an enzyme found only in fish embryos has been identified for the first time, according to a study published in the Journal of Biological Chemistry. Researchers at RIKEN's Glycometabolic Biochemistry Laboratory in Japan named the gene ngly2, and used cryo-electron microscopy to determine the three-dimensional molecular structure of the enzyme it produces.
The enzyme belongs to a class called peptide:N-glycanases, or PNGases, which remove glycans from proteins. Glycans are complex sugar molecules involved in a wide range of biological processes, including protein quality control, cell differentiation, and immune responses. PNGases were first discovered in bacteria and plants, and later found in animals as well.
The first animal PNGase was identified in the eggs and embryos of medaka fish. It turned out that fish carry two distinct types of the enzyme: one active in embryos at acidic pH levels, and another present throughout a fish's entire lifecycle that operates at neutral pH. The neutral version received the lion's share of scientific attention because it corresponds closely to an enzyme called NGLY1 found in humans and other mammals. The acid version, active only in embryos, was largely set aside.
"As acid PNGase appears to be fish specific, not much attention has been paid to this enzyme," said Tadashi Suzuki of the RIKEN Glycometabolic Biochemistry Laboratory. "But we believe that to understand the biology of fish, we have to dig deeper into fish-specific events."
Suzuki and colleague Akinobu Honda led a team that set out to close that gap. Using the structural data from cryo-electron microscopy, the team designed a series of biochemical experiments to test how the enzyme actually functions.
"Formulating hypotheses from structural information and testing them experimentally was particularly fascinating," Honda said. "I was especially excited when the experimental results were consistent with the hypotheses we proposed."
One finding came as a surprise. When the team knocked out ngly2 in zebrafish, the embryos showed little visible effect. The researchers suspect the lack of a clear outcome may be linked to the fact that zebrafish live in freshwater rather than saltwater.
"We have a sneaking suspicion that it may have been difficult to see much of an effect because zebrafish are freshwater fish," Honda said. "But knocking out ngly2 in saltwater fish may have more serious consequences."
The implications of the discovery may extend well beyond fish. The ngly2 gene appears to be conserved across a wide range of aquatic animals, including octopuses, ascidians, sea urchins, corals, and shellfish.
"Since ngly2 is conserved across a broad range of aquatic organisms, including octopuses, ascidians, sea urchins, corals, and shellfish, our observations suggest that this gene may play a role in adaptation to aquatic environments," Suzuki said.
The team's next step, based on Honda's comments, points toward testing the effects of knocking out ngly2 in saltwater species, where the gene's function may be more visible and its role in marine adaptation potentially clearer.
