Related Experiment Video

Updated: Feb 26, 2026

Fully Processed Recombinant KRAS4b: Isolating and Characterizing the Farnesylated and Methylated Protein
07:08

Fully Processed Recombinant KRAS4b: Isolating and Characterizing the Farnesylated and Methylated Protein

Published on: January 16, 2020

6.2K

Structural and functional characterization of a DARPin which inhibits Ras nucleotide exchange

Sandrine Guillard1, Paulina Kolasinska-Zwierz1, Judit Debreczeni2

  • 1Antibody Discovery and Protein Engineering, MedImmune, Milstein Building, Granta Park, Cambridge CB21 6GH, UK.

Nature Communications
|July 15, 2017
PubMed

Insights

Researchers developed DARPin K27, a novel antibody mimetic targeting Ras proteins. This inhibitor shows promise for cancer therapy by blocking Ras nucleotide exchange and downstream signaling, including in mutant Ras forms.

Area of Science:

  • Oncology
  • Molecular Biology
  • Drug Discovery

Background:

  • Ras mutations drive numerous human cancers, yet targeted therapies remain limited.
  • Inhibiting Ras nucleotide exchange is a potential therapeutic strategy requiring further mechanistic understanding.

Purpose of the Study:

  • To describe DARPin K27, an antibody mimetic designed to inhibit Ras nucleotide exchange.
  • To evaluate the efficacy and specificity of K27 as a Ras inhibitor.

Main Methods:

  • Characterization of K27 binding affinity and selectivity for Ras GDP.
  • Intracellular expression of K27 in HCT116 cells.
  • Assessment of downstream signaling, including phosphorylated ERK levels.
  • Evaluation of cell proliferation in soft agar assays.

Main Results:

  • K27 demonstrated high affinity (4 nM Kd) for the inactive Ras GDP form, with selectivity confirmed by structural studies.
  • Intracellular K27 expression reduced active Ras levels and inhibited downstream signaling pathways, notably phosphorylated ERK.
  • K27 treatment slowed the growth of HCT116 cells in soft agar.
  • The inhibitor showed consistent efficacy across various Ras isoforms, including wild-type and oncogenic mutants.

Conclusions:

  • DARPin K27 is a potent, non-covalent inhibitor of Ras nucleotide exchange.
  • K27 effectively reduces active Ras signaling and inhibits cancer cell growth.
  • This antibody mimetic represents a promising new avenue for developing Ras-targeted cancer therapies.

More Related Videos

Comparing the Affinity of GTPase-binding Proteins using Competition Assays10:37

Comparing the Affinity of GTPase-binding Proteins using Competition Assays

Published on: October 8, 2015

9.7K
Genome-wide Protein-protein Interaction Screening by Protein-fragment Complementation Assay PCA in Living Cells08:38

Genome-wide Protein-protein Interaction Screening by Protein-fragment Complementation Assay PCA in Living Cells

Published on: March 3, 2015

14.0K

Related Concept Videos

Small GTPases - Ras and Rho01:24

Small GTPases - Ras and Rho

Ras and Rho are small monomeric GTPases that act downstream of receptor tyrosine kinase (RTK) and regulate various cellular processes. These GTPases switch between active and inactive states by binding to guanine nucleotides.
Three regulatory proteins control their activity:
5.6K
The Ras Gene02:38

The Ras Gene

The Ras-gene-encoded proteins are regulators of signaling pathways controlling cell proliferation, differentiation, or cell survival. The Ras-gene family in humans constitutes three primary members—the HRas, NRas, and KRas. These genes code for four functionally distinct yet closely related proteins—the HRas, NRas, KRas4A, and KRas4B. The involvement of mutant Ras genes in human cancer was first discovered in 1982 and is among the most common causes of human tumorigenesis.
Ras is a...
7.4K
Rab Proteins01:14

Rab Proteins

Rab proteins constitute the largest family of monomeric GTPases, of which 70 members are present in humans. Rab proteins and their effectors regulate consecutive stages of vesicle transport such as vesicle transport, docking, and fusion to the correct recipient membrane.
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...
5.3K
Directionality of Nuclear Transport01:42

Directionality of Nuclear Transport

Ras-related nuclear protein or Ran is a small G protein that cycles between its GTP and GDP bound states. Ran specific regulators, a Ran GTPase Activating Protein or RanGAP present in the cytosol and a Ran guanine nucleotide exchange factor or RanGEF present inside the nucleus regulate GTP/GDP exchange. A high concentration of GTP inside the cells, in addition to this asymmetric distribution of  Ran-specific regulators, leads to a higher RanGTP concentration inside the nucleus. This...
4.7K