To survive and hunt in subzero temperatures, spiders of the genus Clubiona have developed unique antifreeze proteins. A study of arthropods collected in pear orchards near the city of Brno (Czech Republic) enabled scientists to uncover the molecular mechanism that helps these spiders remain active during winter without entering hibernation.

Adaptations to Cold in Spiders

Most spiders cannot survive in cold climates. However, some species, such as Bolyphantes index from Norway, jumping spiders of the genus Philodromus, and species found in Alaska and the Chilean Andes, are able to hunt even in winter. Their resistance to low temperatures is explained by the presence of antifreeze proteins, which partially or completely protect their bodies from freezing.

Unique Proteins of the Clubiona Genus

Recently, scientists have turned their attention to the widespread genus of sac spiders, Clubiona. These spiders produce an antifreeze protein (AFP) with a unique β-solenoid structure, previously found only in polar fish. The protein binds to ice crystals, preventing their growth and stopping bodily fluids from freezing. Despite similarities to proteins synthesized by moths and beetles, Clubiona’s antifreeze proteins evolved independently through convergent evolution—when different organisms develop similar solutions to the same problem.

Experimental Data

Between December 2022 and February 2023, an international team of researchers collected 43 spiders. An extract from a single spider’s body lowered the freezing point of water by more than four degrees Celsius. Even after being diluted twentyfold, this effect persisted, and the growth of ice crystals stopped. The results of the study were published in the FEBS Journal.

Chemical Analysis and Protein Structure

The proteins were purified using five cycles of ice-affinity purification—a method in which proteins freeze together with the solution. Mass spectrometry revealed that AFP exists in several isoforms (registered in GenBank: Isoform 1 — PQ846958, Isoform 2 — PQ846959) and contains glycosylated regions that likely increase the protein’s stability at low temperatures.

Genetic Features

Transcriptomic analysis of active genes identified at least three Clubiona species in the sample: C. pallidula, C. lutescens, and C. brevipes. The sequences of the antifreeze proteins proved to be unique to this genus and were not found in other arthropods or insects.

Molecular Model and Effectiveness

The three-dimensional structure of AFP was modeled using the AlphaFold2 system. The β-solenoid fold with a flat surface lined with threonine residues allows the protein to efficiently bind to ice crystals. Clubiona proteins proved to be extremely effective: after repeated purification cycles, they almost completely bound to the frozen fraction, a phenomenon not observed in the antifreeze proteins of polar fish, such as Atlantic cod.

Application Prospects

Understanding the nature of Clubiona’s antifreeze proteins explains the winter activity of these spiders and opens up prospects for the development of new biotechnologies. Analogs of such proteins could be used to protect agricultural crops from freezing, extend the shelf life of food products, and in medicine—for example, in the cryopreservation of cells and tissues.