Calcium Binding of Transglutaminases: A 43Ca NMR Study Combined with Surface Polarity Analysis

• Published in: Journal of biomolecular structure & dynamics Vol. 19; no. 1; pp. 59 - 74
• Main Authors: Ambrus, Attila, Bányai, István, Weiss, Manfred S., Hilgenfeld, Rolf, Keresztessy, Zsolt, Muszbek, László, Fésüs, László
• Format: Journal Article
• Language: English
• Published: England Taylor & Francis Group 01.08.2001
• Subjects: Amino Acid Sequence; Binding Sites; Calcium - metabolism; Calcium-Binding Proteins - chemistry; Calcium-Binding Proteins - genetics; Calcium-Binding Proteins - metabolism; Factor XIII - chemistry; Factor XIII - genetics; Factor XIII - metabolism; Humans; In Vitro Techniques; Magnetic Resonance Spectroscopy; Models, Molecular; Molecular Sequence Data; Protein Conformation; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Sequence Homology, Amino Acid; Static Electricity; Transglutaminases - chemistry; Transglutaminases - genetics; Transglutaminases - metabolism
• ISSN: 0739-1102; 1538-0254
• DOI: 10.1080/07391102.2001.10506720

Transglutaminases (TGases) form cross-links between glutamine and lysine side-chains of polypeptides in a Ca 2+ -dependent reaction. The structural basis of the Ca 2+ -effect is poorly defined. 43 Ca NMR, surface polarity analysis combined with multiple sequence alignment and the construction of a new homology model of human tissue transglutaminase (tTGase) were used to obtain structural information about Ca 2+ binding properties of factor XIII-A 2 , tTGase and TGase 3 (each of human origin). 43 Ca NMR provided higher average dissociation constants titrating on a wide Ca 2+ -concentration scale than previous studies with equilibrium dialysis performed in shorter ranges. These results suggest the existence of low affinity Ca 2+ binding sites on both FXIII-A and tTGase in addition to high affinity ones in accordance with our surface polarity analysis identifying high numbers of negatively charged clusters. Upon increasing the salt concentration or activating with thrombin, FXIII-A 2 partially lost its original Ca 2+ affinity; the NMR data suggested different mechanisms for the two activation processes. The NMR provided structural evidence of GTP-induced conformational changes on the tTGase molecule diminishing all of its Ca 2+ binding sites. NMR data on the Ca 2+ binding properties of the TGase 3 are presented here; it binds Ca 2+ the most tightly, which is weakened after its proteolytic activation. The investigated TGases seem to have very symmetric Ca 2+ binding sites and no EF-hand motifs.