# Metal Ion Transport by the Cation Diffusion Facilitator Family

> **NIH NIH R01** · NEW YORK UNIVERSITY SCHOOL OF MEDICINE · 2020 · $26,419

## Abstract

This application is for an administrative supplement to parent grant R01 GM125081 to request an instrument
for conducting transport assays in an efficient and automated manner. The SURFE2R N1 from Nanion
Technologies is the first commercial instrument to implement a Solid Supported Membrane (SSM) with
capacitive coupling for measurement of transport from reconstituted proteoliposomes. In our case, these
proteoliposomes will contain YiiP, which is the subject of the parent grant. Transport assays with the SURFE2R
N1 will complement in vitro binding assays to characterize site-specific mutants of YiiP in order to address
mechanistic questions posed in the parent grant application: i.e., the roles of the three different Zn2+ binding
sites on the transport mechanism and on the energetic coupling that employs the proton-motive force to drive
Zn2+ transport. Specifically, this machine will allow us to evaluate coupling stoichiometry (protons transported
per Zn2+ ion) and the effects of site-specific mutations on this stoichiometry. The SURFE2R N1 uses SSM
technology to overcome the low flux rate innate to secondary transporters by adsorbing large numbers of
molecules to the relatively large (3mm) sensor and by using capacitive coupling to measure their combined ion
flow via high-gain/low-noise amplifiers routinely employed for patch-clamp technologies. Very low amounts of
protein are required (0.1-1g) and automation of microfluidic apparatus provides not only for high time
resolution, but also for programmed series of experiments that run without user intervention. Once loaded with
proteoliposomes, a given sensor can be used for up to 100 individual experiments. Thus, this machine
provides a huge performance gain compared to our current protocol that relies on a conventional stop-flow
fluorimeter. In particular, our current protocol requires 10x more protein and does not allow for changes in
proteoliposome buffer conditions. In addition, the availability of real-time data from the SURFE2R N1 with high
time resolution allows fast binding kinetics to be resolved and routine determination of rate constants, EC50 and
IC50 using data analysis software included with the SURFE2R N1, which is far more sophisticated than the
home-made analysis software currently used to analyze data from the fluorimeter. The utility of the SURFE2R
N1, which was not available at the time of the original application, has been validated on over 100 diverse
targets, most of which are either pumps or secondary transporters. Our samples of YiiP were tested at the
Nanion factory, producing robust signals that reflected both steady state transport and pre-steady-state binding
events. As a result, an application for loan of this instrument was approved and Nanion recently shipped a
brand new SURFE2R N1 to our laboratory, where it is awaiting installation. This unit would be available for
immediate purchase should this request be approved. The SURFE2R N1 is also highly effective in quan...

## Key facts

- **NIH application ID:** 10134048
- **Project number:** 3R01GM125081-02S1
- **Recipient organization:** NEW YORK UNIVERSITY SCHOOL OF MEDICINE
- **Principal Investigator:** David L. Stokes
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $26,419
- **Award type:** 3
- **Project period:** 2019-01-01 → 2022-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10134048

## Citation

> US National Institutes of Health, RePORTER application 10134048, Metal Ion Transport by the Cation Diffusion Facilitator Family (3R01GM125081-02S1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10134048. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*