TY - JOUR

T1 - Overproduction of Human Zip (SLC39) Zinc Transporters in Saccharomyces cerevisiae for Biophysical Characterization

AU - Becares, Eva Ramos

AU - Pedersen, Per Amstrup

AU - Gourdon, Pontus

AU - Gotfryd, Kamil

PY - 2021

Y1 - 2021

N2 - Zinc constitutes the second most abundant transition metal in the human body, and it is implicated in numerous cellular processes, including cell division, DNA and protein synthesis as well as for the catalytic activity of many enzymes. Two major membrane protein families facilitate zinc homeostasis in the animal kingdom, i.e., Zrt/Irt-like proteins (ZIPs aka solute carrier 39, SLC39, family) and Zn transporters (ZnTs), essentially conducting zinc flux in the opposite directions. Human ZIPs (hZIPs) regulate import of extracellular zinc to the cytosol, being critical in preventing overaccumulation of this potentially toxic metal, and crucial for diverse physiological and pathological processes, including development of neurodegenerative disorders and several cancers. To date, our understanding of structure-function relationships governing hZIP-mediated zinc transport mechanism is scarce, mainly due to the notorious difficulty in overproduction of these proteins for biophysical characterization. Here we describe employment of a Saccharomyces cerevisiae-based platform for heterologous expression of hZIPs. We demonstrate that yeast is able to produce four full-length hZIP members belonging to three different subfamilies. One target (hZIP1) is purified in the high quantity and homogeneity required for the downstream biochemical analysis. Our work demonstrates the potential of the described production system for future structural and functional studies of hZIP transporters.

AB - Zinc constitutes the second most abundant transition metal in the human body, and it is implicated in numerous cellular processes, including cell division, DNA and protein synthesis as well as for the catalytic activity of many enzymes. Two major membrane protein families facilitate zinc homeostasis in the animal kingdom, i.e., Zrt/Irt-like proteins (ZIPs aka solute carrier 39, SLC39, family) and Zn transporters (ZnTs), essentially conducting zinc flux in the opposite directions. Human ZIPs (hZIPs) regulate import of extracellular zinc to the cytosol, being critical in preventing overaccumulation of this potentially toxic metal, and crucial for diverse physiological and pathological processes, including development of neurodegenerative disorders and several cancers. To date, our understanding of structure-function relationships governing hZIP-mediated zinc transport mechanism is scarce, mainly due to the notorious difficulty in overproduction of these proteins for biophysical characterization. Here we describe employment of a Saccharomyces cerevisiae-based platform for heterologous expression of hZIPs. We demonstrate that yeast is able to produce four full-length hZIP members belonging to three different subfamilies. One target (hZIP1) is purified in the high quantity and homogeneity required for the downstream biochemical analysis. Our work demonstrates the potential of the described production system for future structural and functional studies of hZIP transporters.

KW - family

KW - membrane proteins

KW - overproduction

KW - production platform

KW - protein purification

KW - Saccharomyces cerevisiae

KW - SLC39

KW - solute carrier 39

KW - yeast

KW - zinc

KW - zinc transporters

KW - ZIPs

U2 - 10.3390/cells10020213

DO - 10.3390/cells10020213

M3 - Journal article

C2 - 33494457

AN - SCOPUS:85100483801

VL - 10

JO - Cells

JF - Cells

SN - 2073-4409

IS - 2

M1 - 213

ER -