Isoforms of an N-acetyl-beta-D-glucosaminidase from the Antarctic krill, Euphausia superba: purification and antibody production.
Two forms of the chitinolytic enzyme N-acetyl-beta-D-glucosaminidase (NAGase, EC 3.2.1.52) have been isolated from the Antarctic krill, Euphausia superba, in order to study their potential role in temperature adaptation processes. A chromatographic protocol was developed that allowed complete separation of the two enzyme forms, named NAGase B and NAGase C. The latter was purified to homogeneity with 600-fold enrichment and a yield of 17%. The molecular mass was 150 kDa. NAGase B showed characteristics of a glycoprotein due to affinity towards concanavalin A sepharose, while NAGase C did not. Highly specific polyclonal antibodies to NAGase C [anti-(E. superba-NAGase C)-IgG] showed only negligible cross-reactivity with NAGase B isoforms. A comparison with the Northern krill, Meganyctiphanes norvegica, revealed a corresponding chromatographic pattern with two main activity peaks, for differentiation named NAGase II and NAGase III. Application of the antibody on M. norvegica revealed a high specificity toward NAGase III and a low cross-reactivity with NAGase II. First indication is given that the two forms are no isoenzymes in a strict sense but instead may have different functions in the metabolism of krill.
Peters
G
G
Institut für Meereskunde, Abteilung Zoologie, Kiel, Germany. gpeters@ifm.uni-kiel.de
Saborowski
R
R
Mentlein
R
R
Buchholz
F
F
D000118:Acetylglucosaminidase / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D000818:Animals; D000917:Antibody Formation; D002845:Chromatography; D003429:Cross Reactions; D003445:Crustacea / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D007527:Isoenzymes / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D008970:Molecular Weight; D011817:Rabbits; D013045:Species Specificity
D007527:Isoenzymes; D000118:Acetylglucosaminidase