Introduction of extended LEC14-type branching into core-fucosylated biantennary N-glycan.
A series of enzymatic substitutions modifies the basic structure of complex-type biantennary N-glycans. Among them, a beta1,2-linked N-acetylglucosamine residue is introduced to the central mannose moiety of the core-fucosylated oligosaccharide by N-acetylglucosaminyltransferase VII. This so-called LEC14 epitope can undergo galactosylation at the beta1,2-linked N-acetylglucosamine residue. Guided by the hypothesis that structural modifications in the N-glycan alter its capacity to serve as ligand for lectins, we prepared a neoglycoprotein with the extended LEC14 N-glycan and tested its properties in three different assays. In order to allow comparison to previous results on other types of biantennary N-glycans the functionalization of the glycans for coupling and assay conditions were deliberately kept constant. Compared to the core-fucosylated N-glycan no significant change in affinity was seen when testing three galactoside-specific proteins. However, cell positivity in flow cytofluorimetry was enhanced in six of eight human tumor lines. Analysis of biodistribution in tumor-bearing mice revealed an increase of blood clearance by about 40%, yielding a favorable tumor/blood ratio. Thus, the extended LEC14 motif affects binding properties to cellular lectins on cell surfaces and organs when compared to the core-fucosylated biantennary N-glycan. The results argue in favor of the concept of viewing substitutions as molecular switches for lectin-binding affinity. Moreover, they have potential relevance for glycoengineering of reagents in tumor imaging.
André
Sabine
S
Institut für Physiologische Chemie, Tierärztliche Fakultät, Ludwig-Maximilians-Universität München, Germany. Sabine.Andre@lmu.de
Kojima
Shuji
S
Prahl
Ingo
I
Lensch
Martin
M
Unverzagt
Carlo
C
Gabius
Hans-Joachim
HJ
D000373:Agglutinins / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D002240:Carbohydrate Sequence; D002460:Cell Line; D045744:Cell Line, Tumor; D000939:Epitopes / Q000737:chemistry / Q000378:metabolism; D005434:Flow Cytometry; D005643:Fucose / Q000378:metabolism*; D006023:Glycoproteins / Q000097:blood / Q000138:chemical synthesis / Q000378:metabolism / Q000493:pharmacokinetics; D006031:Glycosylation; D006801:Humans; D037102:Lectins / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011134:Polysaccharides / Q000097:blood / Q000737:chemistry* / Q000378:metabolism* / Q000493:pharmacokinetics; D011485:Protein Binding; D014018:Tissue Distribution
D000373:Agglutinins; D000939:Epitopes; D006023:Glycoproteins; D037102:Lectins; D011134:Polysaccharides; D005643:Fucose