Ras (G12D Mutant) Antibody [C11A12]

Application: Reactivity: Human, Mouse

Usage Information

Dilution
WB1:500-1:3000 IHC1:25-1:100

Application
WB, IHC

Reactivity
Human, Mouse

Source
Rabbit Monoclonal Antibody

Storage Buffer
PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3

Storage (from the date of receipt)
-20°C (avoid freeze-thaw cycles), 2 years

Predicted MW Observed MW
21 kDa 22 kDa
^*Why do the predicted and actual molecular weights differ? The following reasons may explain differences between the predicted and actual protein molecular weight. Post-translational modifications(e.g., phosphorylation, glycosylation); Splice variants and isoforms; Relative charge; Multimerization.

Datasheet & SDS

Biological Description

Specificity
Ras (G12D Mutant) Antibody [C11A12] detects endogenous levels of total Ras (G12D Mutant) protein.

Clone
C11A12

Synonym(s)
GTPase Kras, K-Ras 2, Ki-Ras, c-K-ras, c-Ki-ras, KRAS, KRAS2, RASK2

Background
Ras G12D represents an oncogenic mutant form of the small GTPase K‑Ras, a member of the RAS family of guanine nucleotide–binding switches that regulate mitogenic and survival signaling downstream of receptor tyrosine kinases and other upstream cues. The mutation replaces glycine with aspartate at position 12 within the P‑loop of the G‑domain, in close proximity to the nucleotide and catalytic machinery, and alters local side‑chain packing, hydrogen‑bonding, and electrostatic environment so that the catalytic site adopts conformations that no longer support efficient GTP hydrolysis in the presence of GTPase‑activating proteins. The mutant protein maintains the canonical RAS fold with a G‑domain that binds GTP or GDP and a flexible switch I and switch II region that mediate effector recognition, yet the G12D substitution increases the occupancy of active‑state conformations and disrupts key contacts required for transition state stabilization, which stabilizes the GTP‑bound signaling‑competent form and reduces responsiveness to GAP‑catalyzed inactivation. The activated G12D mutant engages Raf kinases through the RAS‑binding domain, initiating the Raf–MEK–ERK cascade that drives transcriptional programs for proliferation and survival, and at the same time couples to PI3K through direct interaction, supporting the PI3K–AKT axis that promotes growth and cell survival. Additional effector pathways that respond to RAS G12D include Ral guanine nucleotide exchange factors and other downstream nodes, providing multiple parallel outputs that reinforce each other to maintain a proliferative, anti‑apoptotic state. Structural analyses of K‑Ras G12D complexes show that the aspartate side chain at position 12 introduces steric and electrostatic clashes near the γ‑phosphate of GTP and the arginine finger of GAPs, explaining the impaired catalytic geometry while preserving high affinity for GTP and GDP, so that the protein cycles poorly back to the GDP‑bound inactive state and accumulates in an on state that is competent for effector binding. This mutant signaling behavior sustains chronic activation of MAPK and PI3K pathways, which increases cell cycle entry, resistance to apoptotic cues, and altered differentiation patterns, and positions KRAS G12D as a dominant driver of tumorigenesis in tissues where RAS pathway input controls epithelial homeostasis. Endogenous expression of oncogenic K‑Ras G12D stimulates proliferation of epithelial cells and expands progenitor compartments through combined input from Raf–MEK–ERK and PI3K–AKT cascades, without requiring additional upstream receptor hyperactivation, and this property distinguishes the mutant from wild‑type RAS, which depends on tightly regulated growth factor signals.

Background

Ras G12D represents an oncogenic mutant form of the small GTPase K‑Ras, a member of the RAS family of guanine nucleotide–binding switches that regulate mitogenic and survival signaling downstream of receptor tyrosine kinases and other upstream cues. The mutation replaces glycine with aspartate at position 12 within the P‑loop of the G‑domain, in close proximity to the nucleotide and catalytic machinery, and alters local side‑chain packing, hydrogen‑bonding, and electrostatic environment so that the catalytic site adopts conformations that no longer support efficient GTP hydrolysis in the presence of GTPase‑activating proteins. The mutant protein maintains the canonical RAS fold with a G‑domain that binds GTP or GDP and a flexible switch I and switch II region that mediate effector recognition, yet the G12D substitution increases the occupancy of active‑state conformations and disrupts key contacts required for transition state stabilization, which stabilizes the GTP‑bound signaling‑competent form and reduces responsiveness to GAP‑catalyzed inactivation. The activated G12D mutant engages Raf kinases through the RAS‑binding domain, initiating the Raf–MEK–ERK cascade that drives transcriptional programs for proliferation and survival, and at the same time couples to PI3K through direct interaction, supporting the PI3K–AKT axis that promotes growth and cell survival. Additional effector pathways that respond to RAS G12D include Ral guanine nucleotide exchange factors and other downstream nodes, providing multiple parallel outputs that reinforce each other to maintain a proliferative, anti‑apoptotic state. Structural analyses of K‑Ras G12D complexes show that the aspartate side chain at position 12 introduces steric and electrostatic clashes near the γ‑phosphate of GTP and the arginine finger of GAPs, explaining the impaired catalytic geometry while preserving high affinity for GTP and GDP, so that the protein cycles poorly back to the GDP‑bound inactive state and accumulates in an on state that is competent for effector binding. This mutant signaling behavior sustains chronic activation of MAPK and PI3K pathways, which increases cell cycle entry, resistance to apoptotic cues, and altered differentiation patterns, and positions KRAS G12D as a dominant driver of tumorigenesis in tissues where RAS pathway input controls epithelial homeostasis. Endogenous expression of oncogenic K‑Ras G12D stimulates proliferation of epithelial cells and expands progenitor compartments through combined input from Raf–MEK–ERK and PI3K–AKT cascades, without requiring additional upstream receptor hyperactivation, and this property distinguishes the mutant from wild‑type RAS, which depends on tightly regulated growth factor signals.

References
https://pubmed.ncbi.nlm.nih.gov/26902995/ https://pubmed.ncbi.nlm.nih.gov/28740607/

Protein Size (kDa)	PVDF Transfer Membrane Pore Size (μm)
< 10	0.22
10-20	0.22
20-30	0.45
30-45	0.45
45-70	0.45
80-100	0.45
100-140	0.45
> 140	0.45

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Protein Size (kDa)	Separating Gel Percentage
4-40	20%
12-45	15%
10-70	12.5%
15-100	10%
25-100	8%
60-210	5%