Structure of Human Hyaluronidase-1, a Hyaluronan Hydrolyzing Enzyme Involved in Tumor Growth and Angiogenesis^,

Abstract

Mammalian hyaluronidases hydrolyze hyaluronan, a polysaccharide of diverse physiological roles found in all tissues and body fluids. In addition to its function in normal cellular hyaluronan turnover, human hyaluronidase-1 is implicated in cancer proliferation, angiogenesis, and inflammatory diseases; its expression is up-regulated in advanced stages of bladder cancer, whereas the expression of the alternative splice-variants is down-regulated. The crystal structure reveals a molecule composed of two closely associated domains: a catalytic domain that adopts a distorted ( /R) 8 barrel resembling that of bee venom hyaluronidase, and a novel, EGF-like domain, characteristic of involvement in protein-protein interactions and regulatory processes. The structure shows that the fold of this unique EGF-like domain is intact in four alternative splice-variants, whereas the catalytic domain is likely to be unfolded. Thus, these variants may function by competing with the full-length enzyme for the putative protein partner and regulating enzymatic activity in healthy cells.Hyaluronan (HA 1 ) is a linear, unsulfated, negatively charged, glycosaminoglycan formed from ∼2000-25000 repeating disaccharide units of D-glucuronic acid (GlcUA) and N-acetyl-D-glucosamine (GlcNAc) with [GlcUA-( 1f3)-GlcNAc-( 1f4)] n linkages. HA is the major component of cartilage and serves as a joint lubricant. It controls water homeostasis in tissues and the extracellular matrix, which affects cell motility and the distribution and transport of plasma proteins. HA is also implicated in cell proliferation, differentiation, cell-cell recognition, tumor growth and invasion, angiogenesis, and inflammatory responses (1-3). The cellular role of HA and the HA-mediated signal transduction pathways depends on its size (recently reviewed by Stern et al. (4)). Large HA polymers function in organizing the extracellular matrix and serve as lubricant and shock absorber. In shock, septicemia, post surgery, blood loss, and burns, the level of circulating high molecular mass HA increases. These HA polymers are antiangiogenic, immunosuppressive, and anti-inflammatory. In contrast, HA fragments of intermediate sizes are involved in the body's alarm system. They stimulate angiogenesis and inflammatory reactions and facilitate cancer progression and invasion (4). Such HA fragments activate several cytoplasmic and extracellular signal transduction pathways associated with Raf-1 kinase, MAP kinase, and ERK. Short HA oligosaccharides are antiapoptotic and inducers of heat shock proteins. They regulate a different set of signaling molecules than the intermediate size HA including Erb2, PTEN phosphatase, and PI 3 kinase (4).Mammalian hyaluronidases are endo--N-acetyl-hexosaminidases (EC 3.2

Translational research into vascular wall function: regulatory effects of systemic and specific factors

“…Human hyaluronidase has a Ser-353 -Trp-435 C-terminal domain homologous to epidermal growth factor [71] which is present in our model of BTH ( Figure 5) and bee venom hyaluronidases. Epidermal growth factor domain has three disulphide bonds, is essentially stable, regulates binding with other proteins and modulates the activity of a full-size enzyme [70,71].…”

“…However, scarce information about the structure of these biocatalysts hampers the research into regulation of the glycocalyx state. To solve this issue a 3D-model of bovine testicular hyaluronidase (BTH) [70] based on established tertiary structure of human hyaluronidase [71] was constructed using a molecular homological modeling method in silico ( Figure 5). Superposition of BTH with human and bee venom hyaluronidases has revealed differences between these enzymes [70].…”

“…Epidermal growth factor domain has three disulphide bonds, is essentially stable, regulates binding with other proteins and modulates the activity of a full-size enzyme [70,71]. A 3-D structural model of chondroitin-modified BTH was constructed by modeling covalent binding between lysine residues of the enzyme and benzoquinoneactivated chondroitin sulfate links ( Figure 6).…”

Designed Human Serum Hyaluronidase 1 Variant, HYAL1ΔL, Exhibits Activity up to pH 5.9

“…The crystal structures of hyaluronidase from bee (7), wasp (8), and only recently that of human serum hyaluronidase 1 (HYAL1) (9) have been deciphered. In addition to the N-terminal catalytic domain of the insect enzymes, which resembles a distorted (␤/␣) 8 barrel, HYAL1 contains yet another domain.…”

Effect of anticoagulants on the plasma hyaluronidase activities

“…Hyal-1 is the only enzyme present in the circulatory system and is responsible for the catabolism of intracellular and extracellular HA. In contrast, the enzymes involved in the HA synthesis do not circulate freely (2,13). Moreover, augmented expressions of Hyal-1 mRNA and elevated levels of hHyal-1 are documented in several diseases (14)(15)(16)(17).…”