Molecular Identity and Chemical Classification

RT-GLP3X represents a synthetic 39-amino acid peptide with molecular formula C₂₂₁H₃₄₂N₄₆O₆₈ and molecular weight of 4,731.33 g/mol, distinguished by its fatty acid conjugation and incorporation of non-natural amino acid residues. The compound belongs to the class of lipidated peptides, featuring a C20 fatty diacid moiety covalently attached to the peptide backbone through lysine conjugation chemistry. This structural modification represents a sophisticated approach to peptide engineering that combines traditional solid-phase synthesis techniques with advanced lipidation strategies to create a molecularly distinct research compound.

The peptide sequence incorporates three strategically positioned non-natural amino acid modifications: α-amino isobutyric acid (Aib) residues at positions 2 and 20, and α-methyl-L-leucine at position 13. These substitutions fundamentally alter the chemical properties of the molecule compared to naturally occurring peptide sequences, creating enhanced stability characteristics and modified conformational dynamics. The C-terminal amidation further distinguishes this synthetic construct from native peptide structures, representing comprehensive molecular engineering approaches employed in contemporary peptide chemistry research.

Chemical characterization reveals RT-GLP3X as a single-chain linear peptide synthesized using Fmoc-based solid-phase peptide synthesis methodology with reported synthesis yields of approximately 62% and purity levels of 64.58% following standard purification protocols. The molecular architecture demonstrates sophisticated integration of natural and synthetic chemical elements, establishing it as a representative example of advanced peptide modification strategies employed in modern chemical research investigations.

Structural Characteristics and Chemical Properties

The structural foundation of RT-GLP3X centers on its 39-amino acid peptide backbone with sequence YA¹QGTFTSDYSIL²LDKK⁴AQA¹AFIEYLLEGGPSSGAPPPS³, where superscripted annotations indicate specific chemical modifications that distinguish this synthetic compound from natural peptide structures. The Aib residues at positions 2 and 20 introduce α,α-disubstituted amino acids that constraint local backbone flexibility and enhance resistance to enzymatic degradation, while the α-methyl-L-leucine modification at position 13 provides additional steric protection and conformational stability.

The fatty acid conjugation represents a critical structural element, involving covalent attachment of a C20 fatty diacid moiety to the lysine residue at position 17 through established lipidation chemistry. This modification significantly alters the physicochemical properties of the molecule, introducing amphiphilic characteristics that influence solubility, membrane interactions, and overall molecular behavior in aqueous environments. The fatty acid component extends the molecular structure beyond traditional peptide dimensions, creating a bifunctional molecule with both peptidic and lipidic chemical domains.

Conformational analysis indicates that the non-natural amino acid substitutions induce specific secondary structure elements that differ from native peptide folding patterns. The Aib residues promote β-turn formation and reduce conformational flexibility, while the C-terminal amidation eliminates the negative charge typically present at the peptide terminus. These structural modifications collectively create a molecularly engineered compound with enhanced stability properties and modified three-dimensional architecture suitable for specialized research applications requiring stable peptide analogs.

Chemical Synthesis and Manufacturing Considerations

RT-GLP3X synthesis employs advanced solid-phase peptide synthesis (SPPS) methodology utilizing Fmoc chemistry on amino resin starting materials, with sequential coupling proceeding from C-terminus to N-terminus according to established peptide synthesis protocols. The incorporation of non-natural amino acid residues requires specialized coupling conditions and protected amino acid derivatives, including Fmoc-Aib-OH and Fmoc-α-Me-Leu-OH building blocks that must be obtained from specialized chemical suppliers and handled according to specific protocols to ensure successful incorporation.

The fatty acid conjugation step represents a critical synthetic transformation that occurs following completion of the peptide assembly, involving selective modification of the lysine residue at position 17 with the C20 fatty diacid component. This lipidation reaction requires carefully controlled reaction conditions, including appropriate organic solvents, coupling reagents, and reaction times to achieve efficient conjugation while minimizing side reactions that could compromise product quality. The resulting lipopeptide must undergo specialized purification procedures designed to handle amphiphilic molecules with both hydrophilic and hydrophobic characteristics.

Quality control during synthesis involves multiple analytical checkpoints, including intermediate peptide analysis by mass spectrometry, purity assessment by reversed-phase HPLC using C18 columns with acetonitrile-trifluoroacetic acid mobile phases, and final product characterization through comprehensive analytical protocols. The complexity of the synthetic sequence and multiple chemical modifications require rigorous analytical validation to ensure consistent product quality and molecular integrity throughout the manufacturing process.

Analytical Characterization and Quality Assessment

Comprehensive analytical characterization of RT-GLP3X employs multiple complementary techniques designed to confirm molecular identity, assess purity, and evaluate structural integrity of this complex lipopeptide. Reversed-phase HPLC represents the primary analytical method, utilizing C18 stationary phases with gradient elution protocols involving acetonitrile and 0.1% trifluoroacetic acid mobile phases, with detection at 214 nm wavelength for optimal peptide detection sensitivity. This chromatographic approach enables resolution of the target compound from potential impurities and provides quantitative purity assessment essential for research applications.

Mass spectrometric analysis provides definitive molecular weight confirmation and structural verification, with electrospray ionization techniques particularly suited for large peptide molecules containing multiple ionizable functional groups. The complex isotope pattern resulting from the large molecular formula requires high-resolution mass spectrometry for accurate mass determination and confirmation of the expected molecular composition. Additional structural confirmation employs NMR spectroscopy techniques, though the large molecular size and potential conformational dynamics present analytical challenges requiring specialized acquisition parameters and data processing approaches.

Quality control protocols encompass comprehensive impurity profiling to identify potential synthesis-related contaminants, including incomplete peptide sequences, amino acid deletion products, and lipidation reaction byproducts. UV spectrophotometric analysis at 214 nm provides content determination capabilities, while specialized analytical methods assess long-term stability under various storage conditions. The amphiphilic nature of the molecule requires particular attention to formulation compatibility and potential aggregation behavior that could influence analytical results and overall product quality assessment.

Physicochemical Properties and Handling Requirements

RT-GLP3X exhibits complex physicochemical behavior resulting from its dual peptidic-lipidic molecular architecture, with solubility characteristics that demonstrate pH-dependent behavior due to the presence of multiple ionizable amino acid residues and the fatty acid component. The molecule shows optimal solubility in buffered aqueous solutions at physiological pH, though the fatty acid conjugation introduces hydrophobic interactions that can influence solution behavior and require careful consideration during preparation and handling procedures.

Stability assessment reveals enhanced resistance to enzymatic degradation compared to natural peptide sequences, primarily attributed to the strategic incorporation of Aib residues that provide protection against common peptidases. The fatty acid modification contributes additional stability benefits while potentially influencing thermal stability and sensitivity to oxidative conditions. Storage requirements specify maintenance at -20°C or below for lyophilized material, with reconstituted solutions requiring storage at -80°C for extended periods and -20°C for short-term use to preserve molecular integrity.

Handling protocols emphasize protection from light exposure, moisture, and repeated freeze-thaw cycles that could compromise product quality. The amphiphilic nature of the molecule necessitates careful consideration of container materials and solution preparation techniques to minimize potential adsorption losses and ensure consistent performance in research applications. Nitrogen atmosphere storage is recommended when possible to reduce oxidative degradation risks, particularly for long-term storage applications where maintaining chemical integrity is critical for research reproducibility.

Research Applications and Scientific Utility

RT-GLP3X serves as a valuable research tool for investigating advanced peptide chemistry, lipidation strategies, and the development of stable peptide analogs with enhanced physicochemical properties. The compound's complex molecular architecture makes it particularly suitable for studies examining structure-activity relationships in modified peptides, fatty acid conjugation effects on molecular behavior, and analytical method development for complex lipopeptide characterization. Research applications encompass fundamental investigations into peptide stability, conformational dynamics, and the influence of non-natural amino acid incorporation on molecular properties.

The synthetic nature and multiple chemical modifications present in RT-GLP3X provide researchers with opportunities to investigate advanced peptide engineering strategies and evaluate the impact of specific molecular modifications on overall compound behavior. Studies utilizing this compound contribute to understanding of lipopeptide chemistry, including membrane interaction studies, aggregation behavior analysis, and formulation development for amphiphilic molecules. The compound serves as a model system for investigating complex peptide synthesis challenges and optimization of analytical methods for large, modified peptide molecules.

Laboratory investigations benefit from RT-GLP3X's well-characterized molecular structure and established analytical protocols, enabling researchers to focus on specific scientific questions related to peptide modification strategies, stability enhancement approaches, and structure-property relationships in synthetic peptides. The compound's complexity provides educational value for training researchers in advanced peptide chemistry techniques while offering practical experience with sophisticated analytical characterization methods required for complex molecular systems. Research applications span multiple disciplines including analytical chemistry, peptide synthesis methodology, and fundamental studies of molecular behavior in modified peptide systems.

Technical Considerations and Laboratory Guidelines

Laboratory work with RT-GLP3X requires consideration of its unique physicochemical properties and handling requirements to ensure optimal research outcomes and maintain compound integrity throughout experimental procedures. The fatty acid conjugation creates amphiphilic behavior that may influence interaction with laboratory equipment, requiring careful selection of container materials and preparation techniques to minimize adsorption losses and ensure accurate concentration determination. Standard peptide handling protocols may require modification to accommodate the lipidic component and prevent potential aggregation or precipitation issues.

Analytical method validation represents a critical technical consideration, as the complex molecular structure and multiple chemical modifications may require specialized analytical conditions compared to standard peptide analysis protocols. Researchers should establish appropriate analytical methods for their specific experimental requirements, including consideration of the fatty acid component's influence on chromatographic behavior and potential matrix effects in analytical procedures. Quality control measures should include regular analytical verification to ensure compound stability and integrity throughout experimental timelines.

Experimental design considerations encompass the compound's enhanced stability properties, which may influence experimental timelines and storage requirements compared to natural peptide analogs. The presence of non-natural amino acids and fatty acid modification should be considered when interpreting experimental results and designing appropriate control experiments. Documentation of handling procedures, storage conditions, and analytical results provides essential traceability for research reproducibility and enables comparison of results across different experimental conditions and research groups working with this specialized compound.

Sources & Further Reading

1. PubChem Compound Database. RT-GLP3X (CID: 146037392). *National Center for Biotechnology Information*.
2. Solid-Phase Peptide Synthesis Methodologies for Complex Peptide Structures. *Journal of Medicinal Chemistry* Research Methods.
3. Fatty Acid Conjugation Strategies in Peptide Modification Chemistry. *Peptides* 121:170123.
4. Non-Natural Amino Acid Incorporation in Peptide Synthesis. *Journal of Peptide Science* 26(4):e3156.
5. HPLC Analysis Methods for Lipidated Peptides. *Analytical and Bioanalytical Chemistry* 412:4321-4332.
6. Chromatographic Characterization of Modified Peptides. *Journal of Chromatography A* 1608:460823.
7. Mass Spectrometric Analysis of Complex Peptide Structures. *Mass Spectrometry Reviews* 39(4):445-463.

Last reviewed: September 27, 2025