Chemical and immunological characterization of the two alpha-N-acetylgalactosaminidases from squid liver.
Based on the inherent alpha-galactosidase activity, squid liver contains two different alpha-N-acetylgalactosaminidases (alpha-GalNAcases): alpha-N-acetylgalactosaminidase I (alpha-GalNAcase I), which typically exhibits the alpha-galactosidase activity and alpha-N-acetylgalactosaminidase II (alpha-GalNAcase II), which is devoid of such activity. The molecular properties of the alpha-GalNAcases that may account for their enzymological differences are as yet unknown. In this study, we have characterized and compared the chemical and immunological properties of alpha-GalNAcase I and alpha-GalNAcase II. Analysis of the N-terminal sequence of the first twenty amino acids revealed the striking homology between alpha-GalNAcase I and alpha-GalNAcase II. Digestion of alpha-GalNAcase I and alpha-GalNAcase II generated the peptide maps that display similarities in peptide pattern, indicating their close relationship in structure. Polyclonal antibodies were generated in rabbits against the purified alpha-GalNAcase I and alpha-GalNAcase II for comparison of the immunological properties. Both Western blot and surface plasmon resonance (SPR) studies showed that the anti-alpha-GalNAcase II antibody reacted with both alpha-GalNAcase I and alpha-GalNAcase II, whereas the anti-alpha-GalNAcase I antibody reacted only with alpha-GalNAcase I, indicating the presence of common as well as unique antigenic determinants on alpha-GalNAcase I and alpha-GalNAcase II. Taken together, these results suggest that alpha-GalNAcase I and alpha-GalNAcase II are closely related with regard to structure and that their nonhomologous domains are possibly responsible for the differences in enzymatic properties.
Sadik
Golam
G
Department of Health Chemistry, Faculty of Pharmaceutical Science, Niigata University of Pharmacy and Applied Life Sciences, Niigata, Japan.
or Rashid
Md Harun
MH
Itoh-Nashida
Tomoko
T
Ishii
Kazutaka
K
Sato
Yoichi
Y
Shiraishi
Takayuki
T
Uda
Yutaka
Y
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D049832:Decapodiformes / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D018604:Epitope Mapping; D000939:Epitopes / Q000276:immunology*; D007527:Isoenzymes; D008099:Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D017386:Sequence Homology, Amino Acid; D020349:Surface Plasmon Resonance; D048809:alpha-N-Acetylgalactosaminidase / Q000737:chemistry* / Q000276:immunology* / Q000302:isolation & purification
D000939:Epitopes; D007527:Isoenzymes; D048809:alpha-N-Acetylgalactosaminidase