title: Structural significance of galectin design: impairment of homodimer stability by linker insertion and partial reversion by ligand presence.
abstract: Lectins translate information encoded in glycan chains of cellular glycoconjugates into bioeffects. The topological presentation of contact sites for cognate sugar binding is a crucial factor toward this end. To dissect the significance of such phylogenetically conserved properties, the design and engineering of non-natural variants are attractive approaches. Here, a homodimeric human lectin, i.e. adhesion/growth-regulatory galectin-1, is converted into a tandem-repeat display by introducing the 33-amino-acid linker of another family member (i.e. galectin-8). The yield of variant was reduced by about a third. This protein had ∼10-fold higher activity in hemagglutination. Nearly complete sequence determination by mass-spectrometric in-source decay and fingerprinting excluded the presence of any modifications. When (1)H-(15)N heteronuclear single-quantum coherence data on the (15)N-labeled variant and wild-type protein were compared, changes in chemical shifts, signal intensities and resonance multiplicities revealed reduction of stability of interfacial contacts between the lectin domains and an increase in inter-domain flexibility. When both binding sites in the variant were loaded with ligand, association of the two carbohydrate recognition domains was enhanced, corroborated by gel filtration. Dynamic changes in the spatial presentation of the two lectin domains in the context of a tandem-repeat display can alter counterreceptor targeting relative to the fixed positions found in the proto-type galectin homodimer.
authors: Institut für Physiologische Chemie, Tierärztliche Fakultät, Ludwig-Maximilians Universität, Veterinär-straße 13, 80539 München, Germany.
authors: Sabine
authors: S
authors: Vértesy
authors: Abteilung für Angewandte Tumorbiologie, Pathologisches Institut, Klinikum der Ruprecht-Karls-Universität, Im Neuenheimer Feld 224, 69120 Heidelberg, Germany.
authors: Malwina
authors: M
authors: Michalak
authors: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota Health Sciences Center, 6-155 Jackson Hall, 321 Church Str., Minneapolis, MN 55455, USA.
authors: Michelle C
authors: MC
authors: Miller
authors: Funktionelle Proteomanalyse, Deutsches Krebsforschungszentrum (DKFZ), Im Neuenheimer Feld 580, 69120 Heidelberg, Germany.
authors: Martina
authors: M
authors: Schnölzer
authors: Institut für Physiologische Chemie, Tierärztliche Fakultät, Ludwig-Maximilians Universität, Veterinär-straße 13, 80539 München, Germany s.andre@tiph.vetmed.uni-muenchen.de juergen.kopitz@med.uni-heidelberg.de mayox001@umn.edu.
authors: Sabine
authors: S
authors: André
authors: Abteilung für Angewandte Tumorbiologie, Pathologisches Institut, Klinikum der Ruprecht-Karls-Universität, Im Neuenheimer Feld 224, 69120 Heidelberg, Germany s.andre@tiph.vetmed.uni-muenchen.de juergen.kopitz@med.uni-heidelberg.de mayox001@umn.edu.
authors: Jürgen
authors: J
authors: Kopitz
authors: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota Health Sciences Center, 6-155 Jackson Hall, 321 Church Str., Minneapolis, MN 55455, USA s.andre@tiph.vetmed.uni-muenchen.de juergen.kopitz@med.uni-heidelberg.de mayox001@umn.edu.
authors: Kevin H
authors: KH
authors: Mayo
authors: Institut für Physiologische Chemie, Tierärztliche Fakultät, Ludwig-Maximilians Universität, Veterinär-straße 13, 80539 München, Germany.
authors: Hans-Joachim
authors: HJ
authors: Gabius
mesh_terms: D000595:Amino Acid Sequence; D037483:Galectin 1 / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015202:Protein Engineering / Q000379:methods*; D055503:Protein Multimerization; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D020080:Tandem Repeat Sequences
chemical_list: D037483:Galectin 1; D008024:Ligands
keywords: hemagglutination; in-source decay; lectin; sugar code