Cryo-electron microscopy structure and potential enzymatic function of human six-transmembrane epithelial antigen of the prostate 1 (STEAP1)

Abstract

<jats:p>Six-transmembrane epithelial antigen of the prostate 1 (STEAP1) is an integral membrane protein that is highly up-regulated on the cell surface of several human cancers, making it a promising therapeutic target to manage these diseases. It shares sequence homology with three enzymes (STEAP2–STEAP4) that catalyze the NADPH-dependent reduction of iron(III). However, STEAP1 lacks an intracellular NADPH-binding domain and does not exhibit cellular ferric reductase activity. Thus, both the molecular function of STEAP1 and its role in cancer progression remain elusive. Here, we present a ∼3.0-Å cryo-EM structure of trimeric human STEAP1 bound to three antigen-binding fragments (Fabs) of the clinically used antibody mAb120.545. The structure revealed that STEAP1 adopts a reductase-like conformation and interacts with the Fabs through its extracellular helices. Enzymatic assays in human cells revealed that STEAP1 promotes iron(III) reduction when fused to the intracellular NADPH-binding domain of its family member STEAP4, suggesting that STEAP1 functions as a ferric reductase in STEAP heterotrimers. Our work provides a foundation for deciphering the molecular mechanisms of STEAP1 and may be useful in the design of new therapeutic strategies to target STEAP1 in cancer.</jats:p>

Enhanced Stability of Detergent-Free Human Native STEAP1 Protein from Neoplastic Prostate Cancer Cells upon an Innovative Isolation Procedure

“…Based on amino-acid sequence, transmembrane topology, and cellular localization, it was hypothesized that STEAP1 may play an important role as a transporter protein being involved in cell communication [ 3 ] and in the stimulation of cell growth by increasing the levels of reactive oxygen species [ 4 ]. The recent cryo-EM structure-function analysis of STEAP1 cloned in Human Embryonic Kidney (HEK) cells and bound to an antibody-fragment (6Y9B, 2.97 Å resolution) revealed a trimeric arrangement supporting its functional role in heterodimeric assembles with other STEAP1 paralogs to recruit and orient intracellular electron-donating substrates to enable transmembrane-electron transport and the reduction of extracellular metal-ion complexes [ 5 , 6 ].…”

Treating Prostate Cancer by Antibody–Drug Conjugates

“…The molecular mechanisms of STEAP1 activity have been recently elucidated. Enzymatic assays in human cells showed that STEAP1 promotes iron(III) reduction when in STEAP heterotrimers with the intracellular NADPH-binding domain of STEAP4, another member of STEAP family [ 45 ]. Knockdown of STEAP1 gene has been correlated with inhibited cell viability and proliferation and enhanced apoptosis in LNCaP prostate cancer line [ 46 ].…”

The Usefulness of STEAP Proteins in Prostate Cancer Clinical Practice

“…These features pointed out the role of STEAP1 in cellular communication and in cell adhesion processes (19,20). However, the recent cryogenic electron microscopy (cryo-EM) structure of STEAP1 transmembrane domain bound to an antigen-binding fragment of an antibody (mAb 120.545) revealed a trimeric arrangement quite distinct from typical ion channels or transporters (21). The proposed unit indicated that STEAP1 is a functional reductase in heterodimers complexes with other STEAP paralogs with a domain-swapped architecture with the intracellular OxRD positioned beneath the TMD of the adjacent promoter (21,22).…”

“…However, the recent cryogenic electron microscopy (cryo-EM) structure of STEAP1 transmembrane domain bound to an antigen-binding fragment of an antibody (mAb 120.545) revealed a trimeric arrangement quite distinct from typical ion channels or transporters (21). The proposed unit indicated that STEAP1 is a functional reductase in heterodimers complexes with other STEAP paralogs with a domain-swapped architecture with the intracellular OxRD positioned beneath the TMD of the adjacent promoter (21,22). This arrangement supports a model in which the heme-binding site recruit and orient intracellular electron-donating substrates bound to an adjacent STEAP2-4 subunit, enabling transmembrane electron transport and the reduction of extracellular metal-ion complexes (18,21).…”

“…The proposed unit indicated that STEAP1 is a functional reductase in heterodimers complexes with other STEAP paralogs with a domain-swapped architecture with the intracellular OxRD positioned beneath the TMD of the adjacent promoter (21,22). This arrangement supports a model in which the heme-binding site recruit and orient intracellular electron-donating substrates bound to an adjacent STEAP2-4 subunit, enabling transmembrane electron transport and the reduction of extracellular metal-ion complexes (18,21). These structural features highlight the usefulness of STEAP1 as a promising therapeutic target and biomarker for cancer and encouraged the development of strategies targeting STEAP1.…”