‫יפתח ניר‬
‫ פרופ’ עמית מלר‬:‫מנחה‬
‫פורות במצב מוצק‬-‫ פענוח חד מולקולרי של מבנה וקטליזת שרשראות יוביקיוטין באמצעות ננו‬:‫נושא המחקר‬
Iftach Nir
Supervisor: Prof. Amit Meller
Research Title: Exploring Ubiquitin Chains Structure and Catalysis at the Single-Molecule level using
Solid-State Nanopores
Abstract:
Single-molecule analysis provides a direct means to unravel and quantify biomolecular
structure and the interactions among the various components in a biological assembly.
Ubiquitin is a small (8.5 kDa) protein that helps to regulate protein homeostasis in eukaryotic
cells. The regulation takes place by Ubiquitination, which is a post-translational modification
where ubiquitin monomers and/or chains are attached to a substrate protein [1]. D ininietete
the cell’s Ubiquitin turnover rate is important in developing biophysical insights for the cell's
function as well as for future drug development and personalized medicine [2]. In this study,
I use Solid-State Nanopore Sensing (NS) to determine Ubiquitin chains structure and catalytic
status at the single molecule level. NS is a simple yet extremely powerful technique for the
characterization of unlabeled biomolecules [3]. Specifically NS has recently used to detect
individual proteins at their native folded state [4]. Here I employ NS for the first time to
quantitatively measure the Di-Ubiquitin catalysis process. I find that NS is a viable method for
determining and quantifying the products of the catalysis. These single molecule results shed
new light on the catalysis as the ratio of product/substrate can be accurately calculated to
evaluate its efficiency and the distributions of these populations can be estimated. Future
work will include detecting single amino acid mutants and differentiation between Ubiquitin
conformations.
References:
1. Hershko A, Ciechanover A, The ubiquitin system for protein degradation. Annu Rev
Biochem. 1992, 61:761-807.
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2. Fischer D, Disease-specific accumulation of mutant ubiquitin as a marker for
proteasomal dysfunction in the brain. The FASEB Journal. 2003, 17: 2014-2024.
3. Dekker C, Solid-state nanopores, Nature Nanotechnology, 2007, 2(4):209-215.
4. Larkin J, High-Bandwidth Protein Analysis Using Solid-State Nanopores. Biophysical
Journal, 2014, 106: 696-704.
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