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Protein Expr Purif ; 221: 106516, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38801985

RESUMO

Galectins are a large and diverse protein family defined by the presence of a carbohydrate recognition domain (CRD) that binds ß-galactosides. They play important roles in early development, tissue regeneration, immune homeostasis, pathogen recognition, and cancer. In many cases, studies that examine galectin biology and the effect of manipulating galectins are aided by, or require the ability to express and purify, specific members of the galectin family. In many cases, E. coli is employed as a heterologous expression system, and galectin expression is induced with isopropyl ß-galactoside (IPTG). Here, we show that galectin-3 recognizes IPTG with micromolar affinity and that as IPTG induces expression, newly synthesized galectin can bind and sequester cytosolic IPTG, potentially repressing further expression. To circumvent this putative inhibitory feedback loop, we utilized an autoinduction protocol that lacks IPTG, leading to significantly increased yields of galectin-3. Much of this work was done within the context of a course-based undergraduate research experience, indicating the ease and reproducibility of the resulting expression and purification protocols.


Assuntos
Escherichia coli , Galectina 3 , Isopropiltiogalactosídeo , Galectina 3/genética , Galectina 3/metabolismo , Galectina 3/biossíntese , Galectina 3/química , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Isopropiltiogalactosídeo/farmacologia , Expressão Gênica , Galectinas/genética , Galectinas/metabolismo , Galectinas/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Sanguíneas/genética , Proteínas Sanguíneas/metabolismo
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