Recombinant expression, purification and preliminary biophysical and structural studies of C-terminal carbohydrate recognition domain from human galectin-4

Autor(es)

Rustiguel, Joane K. ^[1] ; Kumagai, Patricia S. ^[2] ; Dias-Baruffi, Marcelo ^[3] ; Costa-Filho, Antonio J. ^[4] ; Nonato, Maria Cristina ^[1]

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Lab Cristalog Prot, Fac Ciencias Farmaceut Ribeirao Preto, BR-14040903 Ribeirao Preto, SP - Brazil

^[2] Univ Sao Paulo, Inst Fis Sao Carlos, Grp Biofis Mol Sergio Mascarenhas, BR-13563120 Sao Carlos, SP - Brazil

^[3] Univ Sao Paulo, Fac Ciencias Farmaceut Ribeirao Preto, Lab Glicoimunol, BR-14040903 Ribeirao Preto, SP - Brazil

^[4] Univ Sao Paulo, Fac Filosofia Ciencias & Letras Ribeirao Preto, Lab Biofis Mol, BR-14040901 Ribeirao Preto, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: Protein Expression and Purification; v. 118, p. 39-48, FEB 2016.
• Citações Web of Science: 3

Resumo

Galectin-4 (Gal4), a tandem-repeat type galectin, is expressed in healthy epithelium of the gastrointestinal tract. Altered levels of Gal4 expression are associated with different types of cancer, suggesting its usage as a diagnostic marker as well as target for drug development. The functional data available for this class of proteins suggest that the wide spectrum of cellular activities reported for Gal4 relies on distinct glycan specificity and structural characteristics of its two carbohydrate recognition domains. In the present work, two independent constructs for recombinant expression of the C-terminal domain of human galectin-4 (hGal4-CRD2) were developed. His(6)-tagged and untagged recombinant proteins were overexpressed in Escherichia coli, and purified by affinity chromatography followed by gel filtration. Correct folding and activity of hGal4-CRD2 were assessed by circular dichroism and fluorescence spectroscopies, respectively. Diffraction quality crystals were obtained by vapor-diffusion sitting drop setup and the crystal structure of CRD2 was solved by molecular replacement techniques at 1.78 angstrom resolution. Our work describes the development of important experimental tools that will allow further studies in order to correlate structure and binding properties of hGal4-CRD2 and human galectin-4 functional activities. (C) 2015 Elsevier Inc. All rights reserved. (AU)

• Processo FAPESP: 11/21811-1 - Caracterização estrutural e avaliação de aspectos funcionais de galectinas humanas do grupo tandem-repeat
• Beneficiário: Maria Cristina Nonato
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 11/21767-2 - Interações moleculares no mecanismo de ação da galectina-4 humana
• Beneficiário: Patricia Suemy Kumagai
• Modalidade de apoio: Bolsas no Brasil - Doutorado Direto
• Processo FAPESP: 10/17662-8 - Ressonância magnética eletrônica em estudos de estrutura, função e interações de moléculas com interesse biológico
• Beneficiário: Antonio José da Costa Filho
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 10/16153-2 - Caracterização estrutural e avaliação de aspectos funcionais de galectinas humanas do grupo tandem-repeat
• Beneficiário: Joane Kathelen Rustiguel Bonalumi
• Modalidade de apoio: Bolsas no Brasil - Doutorado