Self-assembly of bacteriophage-associated hyaluronate lyase (HYLP2) into an enzymatically active fibrillar film.
The in vitro assembly of a soluble protein into its mature fibrillar form is usually accompanied by loss of its functional activity. Our study is the first demonstration of a natural enzyme (HylP2) retaining its enzymatic activity on conversion from pre-fibril to mature fibril and supports the contention that minor conformational changes in the native folded form of a protein can lead to the formation of a functional fibril. Hyaluronate lyase (HylP2) is a natural enzyme of bacteriophage 10403 of Streptococcus pyogenes. At pH 5.0, the enzyme undergoes partial unfolding localized in its N-terminal domain while the C-terminal domain maintains its folded trimeric conformation. This structural variant of HylP2 retains about 70% enzymatic activity with hyaluronan. It further self-assembles into a fibrillar film in vitro through solvent-exposed nonpolar surfaces and intermolecular beta-sheet formation by the beta-strands in the protein. Interestingly, the mature fibrillar film of HylP2 also retains about 60 and 20% enzymatic activity for hyaluronic acid and chondroitin sulfate, respectively. The possession of broad substrate specificity by the fibrillar form of HylP2 indicates that fluctuations in pH, which do not lead to loss of functionality of HylP2, might assist in bacterial pathogenesis. The formation of fibrillar film-like structure has been observed for the first time among the hyaluronidase enzymes. After acquiring this film-like structure in bacteriophage, HylP2 still retains its enzymatic activity, which establishes that these fibrils are a genuinely acquired protein fold/structure.
Division of Molecular and Structural Biology, Central Drug Research Institute, Lucknow 226001, India.
Parul
P
Mishra
Vinod
V
Bhakuni
D001435:Bacteriophages / Q000201:enzymology*; D002809:Chondroitin Sulfates / Q000737:chemistry; D006820:Hyaluronic Acid / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D011133:Polysaccharide-Lyases / Q000737:chemistry*; D017510:Protein Folding; D020836:Protein Structure, Quaternary / Q000502:physiology; D017433:Protein Structure, Secondary / Q000502:physiology; D013297:Streptococcus pyogenes / Q000821:virology; D013379:Substrate Specificity / Q000502:physiology; D014764:Viral Proteins / Q000737:chemistry*
D014764:Viral Proteins; D006820:Hyaluronic Acid; D002809:Chondroitin Sulfates; D011133:Polysaccharide-Lyases; C100002:hyaluronate lyase