Late embryogenesis abundant (LEA) proteins are a family of hyper-hydrophilic proteins that accumulate in response to cellular dehydration. Originally identified as plant proteins associated with seed desiccation tolerance, LEA proteins have been identified in a wide range of organisms such as invertebrates and microorganisms. LEA proteins are thought to protect proteins and biomembranes under water-deficit conditions. Here, we characterized WCI16, a wheat (Triticum aestivum) protein that belongs to a class of plant proteins of unknown function, and provide evidence that WCI16 shares common features with LEA proteins. WCI16 was induced during cold acclimation in winter wheat. Based on its amino acid sequence, WCI16 is highly hydrophilic, like LEA proteins, despite having no significant sequence similarity to any of the known classes of LEA proteins. Recombinant WCI16 protein was soluble after boiling, and ¹H-nuclear magnetic resonance (NMR) spectroscopy revealed that the structure of WCI16 is random and has no hydrophobic regions. WCI16 exhibited in vitro cryoprotection of the freeze-labile enzyme l-lactate dehydrogenase as well as double-stranded DNA binding activity, suggesting that WCI16 may protect both proteins and DNA during environmental stresses. The biological relevance of these activities was supported by the subcellular localization of a green fluorescent protein (GFP)-fused WCI16 protein in the nucleus and cytoplasm. Heterologous expression of WCI16 in Arabidopsis (Arabidopsis thaliana) plants conferred enhanced freezing tolerance. Taken together, our results indicate that WCI16 represents a novel class of LEA proteins and is involved in freezing tolerance.