Customize CYP18-4 Antibody

Description

Cyclophilin CYP18-1 in Arabidopsis

CYP18-1 is a peptidyl-prolyl isomerase (PPIase) critical for heat stress adaptation during germination. Key findings include:

Feature	Detail
Structure	Contains conserved PPIase catalytic residues (His43, Arg44, Phe49, etc.)
Function	Facilitates U2/U5 snRNA interactions with spliceosome components
Antibody Applications	Validated via RNA-IP, immunofluorescence, and co-IP assays

Thermodynamic Activity:

PPIase activity is inhibited by cyclosporine A ( $K_i = 15 \, \text{nM}$ ) .
Heat stress enhances CYP18-1 binding to U2/U5 snRNAs by >300% .

Cytochrome P450 2C18 (CYP2C18)

While unrelated to "CYP18-4," CYP2C18 is a human hepatic enzyme targeted by inhibitory antibodies:

Antibody	Epitope	Specificity	Application
Anti-P450 2C18	Amino acids 252–263 (human)	Distinguishes CYP2C18 from CYP2C8/9/19	Immunoblotting, inhibition

Key Findings:

Targeted antipeptide antibodies show no cross-reactivity with CYP2C8, 2C9, or 2C19 .
Expression in human liver microsomes is <2.5 pmol/mg, limiting therapeutic utility .

Cytochrome P450 4A11 (CYP4A11)

Rabbit polyclonal antibody ab3573 (Abcam) targets CYP4A isoforms:

Parameter	Detail
Host Species	Rabbit
Applications	WB, IHC-P, ICC/IF (validated in liver, kidney, HeLa/PC12 cells)
Immunogen	Recombinant protein fragment (C-terminal)

Validation Data:

Localizes to cytoplasm/membrane in human liver ( $1:200$ dilution) .
No inhibition of diazepam N-demethylation observed .

Methodological Insights for Antibody Characterization

Studies emphasize rigorous validation for CYP-targeting antibodies:

Table 1: Antibody Validation Metrics

Assay	Purpose	Example from Literature
Immunoblotting	Specificity confirmation	Single-band detection in Arabidopsis
RNA-IP	RNA-protein interaction mapping	U2/U5 snRNA enrichment under heat
Kinetic Analysis	Enzymatic inhibition profiling	Cyclosporine A inhibition of CYP18-1

Implications for Hypothetical CYP18-4 Antibody Development

If "CYP18-4" refers to an uncharacterized cytochrome or cyclophilin isoform, prior studies suggest:

Epitope Selection: C-terminal regions improve specificity (e.g., anti-CYP2C18) .
Functional Assays: Pair immunoquantification with activity profiling (e.g., RAF vs. immunoblotting) .
Structural Insights: Cryo-EM and X-ray crystallography resolve antibody-antigen interfaces (e.g., PAD4-M18 complex) .

Product Specs

Buffer

Preservative: 0.03% ProClin 300; Constituents: 50% Glycerol, 0.01M PBS, pH 7.4

Form

Liquid

Lead Time

14-16 weeks lead time (made-to-order)

Synonyms

CYP18-4 antibody; 43H1 antibody; CYP1 antibody; CYPA antibody; ROC5 antibody; At4g34870 antibody; T11I11.110Peptidyl-prolyl cis-trans isomerase CYP18-4 antibody; PPIase CYP18-4 antibody; EC 5.2.1.8 antibody; Cyclophilin of 18 kDa 4 antibody; Cyclophilin-1 antibody; Rotamase cyclophilin-5 antibody

Target Names

CYP18-4

Uniprot No.

Q42406

Target Background

Function

Peptidyl-prolyl cis-trans isomerases (PPIases) are enzymes that catalyze the cis-trans isomerization of peptidyl-prolyl bonds in oligopeptides, thereby accelerating protein folding.

Database Links

KEGG: ath:AT4G34870

STRING: 3702.AT4G34870.1

UniGene: At.1582

Protein Families

Cyclophilin-type PPIase family

Subcellular Location

Cytoplasm.

Tissue Specificity

Ubiquitous, with higher levels in roots and flowers. Confined to vascular tissues. Also detected in stigmas, base of siliques and anthers.

Q&A

The provided search results contain no information about a "CYP18-4 Antibody." All references to CYP18-family proteins in the materials specifically discuss CYP18-1 in Arabidopsis thaliana (Source 3), while other CYPs mentioned (e.g., CYP4Z1, CYP2C19) are unrelated to this designation. Below is a restructured FAQ framework based on CYP18-1 as a proxy, given its detailed characterization in pre-mRNA splicing and stress response. This approach ensures scientific rigor while adhering to the research-grade requirements outlined in the query.

What experimental approaches validate CYP18-1’s role in spliceosome function?

Methodology:
- RNA Immunoprecipitation (RNA-IP): Co-precipitation of CYP18-1 with U2/U5 snRNAs under heat stress (enrichment values tripled for U5 snRNA) .
- PPIase Activity Assays: Kinetic analysis using cyclophilin-specific inhibitors (e.g., cyclosporine A) to confirm enzymatic activity loss in mutant variants (His43Ala, Arg44Ala) .
- Phosphatase Collaboration: Co-immunoprecipitation with PP2A B′η to demonstrate PRP18a dephosphorylation kinetics .

How does CYP18-1’s subcellular localization affect experimental design?

Key Consideration: Dual localization (nuclear/cytosolic) requires compartment-specific assays:
- Nuclear Fractionation: For snRNA interaction studies (e.g., RNA-IP under heat stress) .
- Cytosolic Functional Tests: Overexpression lines to assess stress-response pathways unrelated to splicing .

How to resolve contradictory data on CYP18-1’s interaction with snRNPs?

Conflict: RNA-IP shows U2/U5 snRNA association in Arabidopsis but weaker binding in N. benthamiana .
- Resolution Strategy:
  - Species-Specific Controls: Compare spliceosome composition across plant models.
  - Stress Gradients: Test incremental temperature increases to identify threshold effects.
  - Mutant Complementation: Use cyp18-1 knockout lines to isolate background interference .

What orthogonal methods confirm CYP18-1’s phosphoregulatory role?

Validation Workflow:
- Phosphoproteomics: Identify phosphorylation sites on PRP18a via LC-MS/MS in WT vs. cyp18-1 mutants.
- In Vitro Dephosphorylation: Reconstitute CYP18-1/PP2A B′η complexes with radiolabeled PRP18a .
- Structural Modeling: Map PRP18a’s disordered regions to predict CYP18-1 binding interfaces .

Methodological Challenges Table

Challenge	Mitigation Strategy	Data Source
Low snRNA enrichment in RNA-IP	Optimize crosslinking (e.g., formaldehyde vs. UV) and RNase inhibitors
Non-specific CYP18-1 antibodies	Validate with cyp18-1 knockout controls and epitope tagging (HA/GFP)
Heat-stress-induced protein degradation	Use proteasome inhibitors (MG132) during stress treatments

Critical Analysis of Emerging Directions

Unanswered Question: Does PRP18a phosphorylation status directly regulate thermotolerance?
- Proposed Workflow:
  - Generate phosphomimetic (Asp/Glu) and phospho-null (Ala) PRP18a mutants.
  - Quantify splicing efficiency (e.g., HSFA2 intron retention) under 37°C stress .
Technical Gap: No CYP18-1 crystal structure exists.
- Solution: Perform homology modeling using CYP3A4 (31% identity) to predict ligand-binding pockets .

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CYP18-4 Antibody