Human NEU3 and NEU4 enzymes prefer short polysialic acid substrates

Regulation of sialic acids is important to many biological processes and involves both biosynthesis and catabolism. The human neuraminidase enzymes (hNEU) regulate many sialosides in human health and disease. Polysialic acid (polySia) is a polyanion of α(2 → 8) linked sialic acids with roles in nervous, reproductive, and immune systems and is dysregulated in some malignancies and mental disorders. The unique chemical properties of this polymer, which include an enhanced susceptibility to acid-catalyzed hydrolysis, have hampered its study. Herein we describe the first systematic study of hNEU isoenzyme activity towards polySia in vitro. Our experimental design allowed us to study the impact of several factors that may influence polysialic acid degradation including pH, polymer size, and the relative ionic strength of the media. We report that short chains of polySia (oligoSia, degree of polymerization, DP 3-8) were substrates of NEU3 and NEU4 at acidic pH, but not at neutral pH. We were unable to detect hNEU-catalyzed hydrolysis of longer polymers (polySia, DP 10-20) over background hydrolysis. These findings suggest that longer polySia oligomers may be degraded through neuraminidase-independent mechanisms, such as internalization or degradation in endosomes and lysosomes.