TY - JOUR

T1 - Isolation and Characterization of Nanobodies against a Zinc-Transporting P-Type ATPase

AU - Longhin, Elena

AU - Gronberg, Christina

AU - Hu, Qiaoxia

AU - Duelli, Annette Susanne

AU - Andersen, Kasper Rojkjaer

AU - Laursen, Nick Stub

AU - Gourdon, Pontus

PY - 2018

Y1 - 2018

N2 - P-type ATPases form a large and ubiquitous superfamily of ion and lipid transporters that use ATP (adenosine triphosphate) to carry out their function. The IB subclass (P-IB-ATPases) allows flux of heavy metals and are key players in metal detoxification, critical for human health, crops, and survival of pathogens. Nevertheless, P-IB-ATPases remain poorly understood at a molecular level. In this study, nanobodies (Nbs) are selected against the zinc-transporting P-IB-ATPase ZntA from Shigella sonnei (SsZntA), aiming at developing tools to assist the characterization of the structure and function of this class of transporters. We identify six different Nbs that bind detergent stabilized SsZntA. We further assess the effect of the Nbs on the catalytic function of SsZntA, and find that five nanobodies associate without affecting the function, while one nanobody significantly reduces the ATPase activity. This study paves the way for more refined mechanistical and structural studies of zinc-transporting P-IB-ATPases.

AB - P-type ATPases form a large and ubiquitous superfamily of ion and lipid transporters that use ATP (adenosine triphosphate) to carry out their function. The IB subclass (P-IB-ATPases) allows flux of heavy metals and are key players in metal detoxification, critical for human health, crops, and survival of pathogens. Nevertheless, P-IB-ATPases remain poorly understood at a molecular level. In this study, nanobodies (Nbs) are selected against the zinc-transporting P-IB-ATPase ZntA from Shigella sonnei (SsZntA), aiming at developing tools to assist the characterization of the structure and function of this class of transporters. We identify six different Nbs that bind detergent stabilized SsZntA. We further assess the effect of the Nbs on the catalytic function of SsZntA, and find that five nanobodies associate without affecting the function, while one nanobody significantly reduces the ATPase activity. This study paves the way for more refined mechanistical and structural studies of zinc-transporting P-IB-ATPases.

KW - P-type ATPase

KW - nanobody

KW - llama

KW - Zinc-transport

KW - Zinc-transporting P-ATPase

KW - ZntA

U2 - 10.3390/antib7040039

DO - 10.3390/antib7040039

M3 - Journal article

C2 - 31544889

VL - 7

JO - Antibodies

JF - Antibodies

SN - 2073-4468

IS - 4

M1 - 39

ER -