Specific Conformation Research Articles

Leveraging Ligand Desymmetrization to Enrich Structural Diversity of Zirconium Metal-Organic Frameworks for Toxic Chemical Adsorption.

The discovery of metal-organic frameworks (MOFs) with novel structures provides significant opportunities for developing porous solids with new properties and enriching the structural diversity of functional materials for various applications. The rational design of building units with specific geometric conformations is essential to direct the construction of MOFs with unique properties. Herein, we leverage a ligand desymmetrization approach to construct a series of new MOFs. A flexible tetratopic carboxylate ligand with a tetrahedral geometry was designed and assembled with a Zr6 cluster, generating four Zr-based MOF structures: NU-2600, NU-2700, NU-2800, and NU-1802, in which the ligand configurations and Zr6 cluster connectivities can be controlled by varying solvents and modulators during synthesis. Except for NU-1802, these represent entirely new topologies. Notably, NU-1802 exhibits structural flexibility, with up to a 74% reduction in the unit cell volume as confirmed by single-crystal X-ray diffraction studies. Given their microporous structures, we studied the adsorption behaviors of n-hexane and 2-chloroethyl ethyl sulfide to explore the structure-property relationships of these MOFs. Overall, this work highlights ligand desymmetrization as a powerful method to enrich MOF structural diversity and access complex MOFs with non-default topologies suitable for applications such as toxic gas capture.

Leveraging Ligand Desymmetrization to Enrich Structural Diversity of Zirconium Metal–Organic Frameworks for Toxic Chemical Adsorption

The discovery of metal–organic frameworks (MOFs) with novel structures provides significant opportunities for developing porous solids with new properties and enriching the structural diversity of functional materials for various applications. The rational design of building units with specific geometric conformations is essential to direct the construction of MOFs with unique properties. Herein, we leverage a ligand desymmetrization approach to construct a series of new MOFs. A flexible tetratopic carboxylate ligand with a tetrahedral geometry was designed and assembled with a Zr6 cluster, generating four Zr‐based MOF structures: NU‐2600, NU‐2700, NU‐2800, and NU‐1802, in which the ligand configurations and Zr6 cluster connectivities can be controlled by varying solvents and modulators during synthesis. Except for NU‐1802, these represent entirely new topologies. Notably, NU‐1802 exhibits structural flexibility, with up to a 74% reduction in the unit cell volume as confirmed by single‐crystal X‐ray diffraction studies. Given their microporous structures, we studied the adsorption behaviors of n‐hexane and 2‐chloroethyl ethyl sulfide to explore the structure‐property relationships of these MOFs. Overall, this work highlights ligand desymmetrization as a powerful method to enrich MOF structural diversity and access complex MOFs with non‐default topologies suitable for applications such as toxic gas capture.

Unveiling the spin adaptability of electrocatalytic nitrogen reduction catalyzed by Fe, Ru and Os-anchored nitrogen-doped graphene

It is of a great challenge to achieve efficient single-atom catalysts (SACs) for electrocatalytic nitrogen reduction. Especially, the SACs from transition metals have complex electron conformation. It is urgently necessary to examine the effect of spin state of transition metals on the catalytic performance of SACs for nitrogen fixed. In this work, using density functional theory (DFT) calculations, we firstly evaluated the catalytic capability of 21 transition metals to achieve the most potential metals with the special electron conformation of d6, including Fe, Ru and Os, to construct SACs. Our examinations revealed that the catalytic performance of SAC for nitrogen reduction increases with the decrease of the spin multiplicity for the same metal through the same reaction pathway and further improves with the increase of period number of transition metal. Additionally, the selectivity of SAC for nitrogen reduction reaction also increases with the increase of period over hydrogen evolution reaction under the low spin state. Finally, we found that the OsN4C catalysts had excellent catalytic performance for nitrogen reduction with the extremely low overpotential (0.03V) through the distal reaction path under the low spin state, which were expected to be the potential industrial applications for electrocatalytic nitrogen reduction.

A conserved mechanism couples cytosolic domain movements to pore gating in the TRPM2 channel

Transient Receptor Potential Melastatin 2 (TRPM2) cation channels contribute to immunocyte activation, insulin secretion, and central thermoregulation. TRPM2 opens upon binding cytosolic Ca2+ and ADP ribose (ADPR). We present here the 2.5 Å cryo-electronmicroscopy structure of TRPM2 from Nematostella vectensis (nvTRPM2) in a lipid nanodisc, complexed with Ca2+ and ADPR-2'-phosphate. Comparison with nvTRPM2 without nucleotide reveals that nucleotide binding-induced movements in the protein's three "core" layers deconvolve into a set of rigid-body rotations conserved from cnidarians to man. By covalently crosslinking engineered cysteine pairs we systematically trap the cytosolic layers in specific conformations and study effects on gate opening/closure. The data show that nucleotide binding in Layer 3 disrupts inhibitory intersubunit interactions, allowing rotation of Layer 2 which in turn expands the gate located in Layer 1. Channels trapped in that "activated" state are no longer nucleotide dependent, but are opened by binding of Ca2+ alone.

Significant Chiral Asymmetry Observed in Neutral Amino Acid Ultraviolet Photolysis.

The origin of homochirality in biological organisms remains an open question. Some suggest that its origin might be extraterrestrial, specifically due to the exposure of chiral molecules to circularly polarized photons in interstellar space, which could cause an initial population imbalance leading to the homochirality observed today. However, this extraterrestrial hypothesis has not been widely accepted, largely due to the belief that molecular optical rotatory dispersion is too insignificant to create the substantial imbalance required for homochirality. Here we report experimental evidence that specific conformers of neutral amino acids exhibit significant asymmetry in the chiral destroying dissociation rate induced by circularly polarized photons. The observed anisotropy factor for the lowest energy conformer of leucine was remarkably large, reaching 0.1─a factor of 13 times larger than observed for zwitterionic leucine in solid films, and nearly 40 times greater than the anisotropy reported in the electronic absorption spectrum of gas-phase leucine ensembles at room temperature. This significant finding indicates that even if reported anisotropy values in the electronic absorption spectrum are low, the dissociation asymmetry of certain conformers can still be substantial. An anisotropy factor of 0.1 could result in an initial enantiomeric excess exceeding 10%, even with a 90% extent of reaction. This discovery suggests that asymmetric photodissociation of amino acids may have been a crucial factor in the emergence of biological homochirality.

8041 Eneboparatide, A Potent Stimulator Of Urinary Calcium Reabsorption, Induces Prolonged Renal Cortex Retention And PTH1 Receptor Signaling

Abstract Disclosure: G. Ravel: Employee; Self; Amolyt Pharma. R. Datta: Consulting Fee; Self; Amolyt Pharma. S. Milano: Employee; Self; Amolyt Pharma. M. Ovize: Employee; Self; Amolyt Pharma. S. Allas: Employee; Self; Amolyt Pharma. M. Aouadi: None. M.D. Culler: Employee; Self; Amolyt Pharma. Natural parathyroid hormone (PTH) replacement therapy for chronic hypoparathyroidism (cHP) is not ideal due to both short circulating half-life and brief receptor signaling, making the effect too transient. Following agonist binding to the PTH 1 receptor (PTH1R), the magnitude and duration of the receptor signal, and consequently the biological response, appears to depend on the affinity of the ligand for a specific receptor conformation termed R0, which determines the duration of ligand binding. Eneboparatide (Enebo) is a hybrid analog of PTH and PTH-related peptide designed to strongly bind to the R0 conformation while having a short circulating half-life similar to that of PTH. When tested in cHP patients, Enebo eliminated the need for calcium (Ca) (88% patients) and Vitamin D (92% patients) supplementation, while maintaining serum Ca levels over a full 24 hours. Urinary calcium (uCa) was rapidly decreased and was normalized in 92% of patients who were hypercalciuric prior to treatment. To explore the mechanism of the potent effect of Enebo on uCa, a binding assay specific for the R0 conformation of the PTH1R confirmed the high affinity of Enebo versus natural PTH1-34, with IC50 = 1.5nM and 30.9nM, respectively. Postulating that the high-affinity binding of Enebo to R0 should result in longer tissue retention in vivo, we examined the tissue levels of PTH1-34 and Enebo in male, 200-300g rats using quantitative whole-body autoradiography (QWBA) following subcutaneous injection of 3H-labeled Enebo and PTH1-34. Tissue levels of the compounds were normalized based on the concentrations observed in cardiac blood. The highest level observed for both compounds was in the renal cortex, the major site of active PTH-induced Ca reabsorption, with peak concentrations observed 30 minutes post-injection. Six hours after injection, 96.3% of PTH1-34 had disappeared from the renal cortex, as compared with only 61.4% of Enebo, despite both compounds having similar circulating half-lives. With respect to activating the PTH1R, both compounds showed a similar increase in cAMP in human embryonic kidney (HEK293) cells transfected with the PTH1R and the Green Downward cADDis cAMP biosensor, with EC50 of 0.32 and 0.48nM for Enebo and PTH1-34, respectively. To mimic the effect of rapid elimination in vivo, the ligands were added to the cells, washed-off after 15 minutes, and cAMP production followed over the next 16 hours. After washout, the cAMP signal induced by PTH1-34 rapidly returned to baseline in less than 60 minutes, whereas the cAMP signal induced by Enebo gradually decreased and was still evident after 16 hours. Collectively, these data confirm high affinity of Enebo for the R0 conformation of the PTH1R, as well as prolonged receptor signaling and retention by the renal cortex, clearly differentiating Enebo from PTH1-34 and providing strong rationale for the efficacy of Enebo in reducing uCa in cHP patients. Presentation: 6/3/2024

A Mathematical Model of Interstitial Fluid Flow and Retinal Tissue Deformation in Macular Edema.

This study aims to develop a mathematical model to elucidate fluid circulation in the retina, focusing on the movement of interstitial fluid (comprising water and albumin) to understand the mechanisms underlying exudative macular edema (EME). The model integrates physiological factors such as retinal pigment epithelium (RPE) pumping, osmotic pressure gradients, and tissue deformation. It accounts for spatial variability in hydraulic conductivity (HC) across the retina and incorporates the structural role of Müller cells (MCs) in maintaining retinal stability. The model predicts that tissue deformation is maximal at the center of the fovea despite fluid exudation from blood capillaries occurring elsewhere, aligning with clinical observations. Additionally, the model suggests that spatial variability in HC across the thickness of the retina plays a protective role against fluid accumulation in the fovea. Despite inherent simplifications and uncertainties in parameter values, this study represents a step toward understanding the pathophysiology of EME. The findings provide insights into the mechanisms underlying fluid dynamics in the retina and fluid accumulation in the foveal region, showing that the specific conformation of Müller cells is likely to play a key role.

N-Glycosylation-Induced Pathologic Protein Conformations as a Tool to Guide the Selection of Biologically Active Small Molecules.

Post-translational modifications such as protein N-glycosylation, significantly influence cellular processes. Dysregulated N-glycosylation, exemplified in Grp94, a member of the Hsp90 family, leads to structural changes and the formation of epichaperomes, contributing to pathologies. Targeting N-glycosylation-induced conformations offers opportunities for developing selective chemical tools and drugs for these pathologic forms of chaperones. We here demonstrate how a specific Grp94 conformation induced by N-glycosylation, identified previously via molecular dynamics simulations, rationalizes the distinct behavior of similar ligands. Integrating dynamic ligand unbinding information with SAR development, we differentiate ligands productively engaging the pathologic Grp94 conformers from those that are not. Additionally, analyzing binding site stereoelectronic properties and QSAR models using cytotoxicity data unveils relationships between chemical, conformational properties, and biological activities. These findings facilitate the design of ligands targeting specific Grp94 conformations induced by abnormal glycosylation, selectively disrupting pathogenic protein networks while sparing normal mechanisms.

Computational Modeling of the Conformation-Dependent Atmospheric Reactivity of Criegee Intermediates.

The impacts of Criegee intermediates (CIs) on atmospheric chemistry depend significantly on the CI conformation. In this Perspective, I highlight examples of how electronic structure and statistical rate theory calculations, in conjunction with experiment, have revealed conformation-dependent details of both CI ground-state reactivity and electronic excitation. Calculations using single-reference electronic structure methods and conventional transition state theory have predicted that CIs with syn-alkyl or syn-vinyl substituents isomerize rapidly to vinyl hydroperoxides (VHPs) or dioxoles, both of which can decompose rapidly under atmospheric conditions. Ongoing computational research on hydroxyl radical (OH) roaming initiated by VHP dissociation requires the application of multireference electronic structure methods and variational transition state theory. CIs that lack both syn-alkyl and syn-vinyl substituents undergo either bimolecular reaction or π* ← π electronic excitation in the atmosphere. Accurate predictions of CI ultraviolet-visible spectra require multireference calculations with large active spaces and at least a second-order perturbative treatment of dynamic electron correlation. The extent to which electronic spectra can be diagnostic of the presence of specific CI conformers varies significantly with CI chemical identity.

Freezing Conformers for Gas-Phase Förster Resonance Energy Transfer.

Various techniques are available to illuminate geometric structures of molecular ions in gas phase, such as Förster Resonance Energy Transfer (FRET) informing on distances between two dyes covalently attached to a molecule. Typically, cationic rhodamines, which absorb and emit visible light, are used for labeling. Extensive work has revealed that the transition energy of a rhodamine is intricately linked to its nearby microenvironment, with nearby charges causing Stark-shifted emission. This occurs because the inter-dye Coulomb interaction is weaker in the excited state (S1) than in the ground state (S0) due to the increase in polarizability upon excitation. Therefore, absorption and emission spectra, along with FRET efficiencies, provide insights into structural motifs. At room temperature, multiple conformers often co-exist, leading to overlapping absorption bands among different conformers and broad spectra. To study specific conformers, it is necessary to isolate them, for example, using ion-mobility spectrometry. Another approach is to reduce temperature, which results in spectral narrowing and distinct absorption bands, allowing for the selection of specific conformers through selective excitation. Here, we describe the instrumentation used for cryogenically cold FRET experiments and discuss recent results for small model systems, as well as future directions for a technique still in its infancy.

Delayed Excitations Induce Polymer Looping and Coherent Motion.

We consider inhomogeneous polymers driven by energy-consuming active processes which encode temporal patterns of athermal kicks. We find that such temporal excitation programs, propagated by tension along the polymer, can effectively couple distinct polymer loci. Consequently, distant loci exhibit correlated motions that fold the polymer into specific conformations, as set by the local actions of the active processes and their distribution along the polymer. Interestingly, active kicks that are canceled out by a time-delayed echo can induce strong compaction of the active polymer.

Phosphorylation patterns in the AT1R C-terminal tail specify distinct downstream signaling pathways.

Different ligands stabilize specific conformations of the angiotensin II type 1 receptor (AT1R) that direct distinct signaling cascades mediated by heterotrimeric G proteins or β-arrestin. These different active conformations are thought to engage distinct intracellular transducers because of differential phosphorylation patterns in the receptor C-terminal tail (the "barcode" hypothesis). Here, we identified the AT1R barcodes for the endogenous agonist AngII, which stimulates both G protein activation and β-arrestin recruitment, and for a synthetic biased agonist that only stimulates β-arrestin recruitment. The endogenous and β-arrestin-biased agonists induced two different ensembles of phosphorylation sites along the C-terminal tail. The phosphorylation of eight serine and threonine residues in the proximal and middle portions of the tail was required for full β-arrestin functionality, whereas phosphorylation of the serine and threonine residues in the distal portion of the tail had little influence on β-arrestin function. Similarly, molecular dynamics simulations showed that the proximal and middle clusters of phosphorylated residues were critical for stable β-arrestin-receptor interactions. These findings demonstrate that ligands that stabilize different receptor conformations induce different phosphorylation clusters in the C-terminal tail as barcodes to evoke distinct receptor-transducer engagement, receptor trafficking, and signaling.

Mechanochemical Diversity in Block Copolymers.

Covalent polymer chains are known to undergo mechanochemical events when subjected to mechanical forces. Such force-coupled reactions, like C-C bond scission in homopolymers, typically occur in a non-selective manner but with a higher probability at the mid-chain. In contrast, block copolymers (BCPs), composed of two or more chemically distinct chains linked by covalent bonds, have recently been shown to exhibit significantly different mechanochemical reactivities and selectivities. These differences may be attributable to the atypical conformations adopted by their chains, compared to the regular random coil. Beyond individual molecules, when BCPs self-assemble into ordered aggregates in solution, the non-covalent interactions between the chains lead to meaningful acceleration in the activation of embedded force-sensitive motifs. Furthermore, the microphase segregation of BCPs in bulk creates periodically dispersed polydomains, locking the blocks in specific conformations which have also been shown to affect their mechanochemical reactivity, with different morphologies influencing reactivity to varying extents. This review summarizes the studies of mechanochemistry in BCPs over the past two decades, from the molecular level to assemblies, and up to bulk materials.

Uncovering a Latent Bioactive Interleukin-6 Glycoform.

A bioinspired semisynthesis of human-interleukin-6 bearing N-glycan at Asn143 (143glycosyl-IL-6) was performed by intentional glycosylation effects and protein folding chemistry for regioselective peptide-backbone activation. 143Glycosyl-IL-6 is a genetically coded cytokine, but isolation was difficult owing to a tiny amount. IL6-polypeptide (1-141-position) with an intentionally inserted cysteine at 142-position was expressed in E. coli. The expressed polypeptide was treated with a chemical folding process to make a specific helices bundle conformation through native two-disulfide bonds (43-49 and 72-82). Utilizing the successfully formed free-142-cysteine, sequential conversions using cyanation of 142-cysteine, hydrazinolysis, and thioesterification created a long polypeptide (1-141)-thioester. However, the resultant polypeptide-thioester caused considerable aggregation owing to a highly hydrophobic peptide sequence. After the reduction of two-disulfide bonds of polypeptide (1-141)-thioester, an unprecedented hydrophilic N-glycan tag was inserted at the resultant cysteine thiols. The N-glycan tags greatly stabilized polypeptide-thioester. The subsequent native chemical ligation and desulfurization successfully gave a whole 143glycosyl-IL-6 polypeptide (183-amino acids). Removal of four N-glycan tags and immediate one-pot in vitro folding protocol efficiently produced the folded 143glycosyl-IL-6. The folded 143glycosyl-IL-6 exhibited potent cell proliferation activity. The combined studies with molecular dynamics simulation, semisynthesis, and bioassays predict the bioactive conformation of latent 143glycosyl-IL-6.

Uncovering a Latent Bioactive Interleukin‐6 Glycoform

A bioinspired semisynthesis of human‐interleukin‐6 bearing N‐glycan at Asn143 (143glycosyl‐IL‐6) was performed by intentional glycosylation effects and protein folding chemistry for regioselective peptide‐backbone activation. 143Glycosyl‐IL‐6 is a genetically coded cytokine, but isolation was difficult owing to a tiny amount. IL6‐polypeptide (1‐141‐position) with an intentionally inserted cysteine at 142‐position was expressed in E. coli. The expressed polypeptide was treated with a chemical folding process to make a specific helices bundle conformation through native two‐disulfide bonds (43–49 and 72–82). Utilizing the successfully formed free‐142‐cysteine, sequential conversions using cyanation of 142‐cysteine, hydrazinolysis, and thioesterification created a long polypeptide (1–141)‐thioester. However, the resultant polypeptide‐thioester caused considerable aggregation owing to a highly hydrophobic peptide sequence. After the reduction of two‐disulfide bonds of polypeptide (1–141)‐thioester, an unprecedented hydrophilic N‐glycan tag was inserted at the resultant cysteine thiols. The N‐glycan tags greatly stabilized polypeptide‐thioester. The subsequent native chemical ligation and desulfurization successfully gave a whole 143glycosyl‐IL‐6 polypeptide (183‐amino acids). Removal of four N‐glycan tags and immediate one‐pot in vitro folding protocol efficiently produced the folded 143glycosyl‐IL‐6. The folded 143glycosyl‐IL‐6 exhibited potent cell proliferation activity. The combined studies with molecular dynamics simulation, semisynthesis, and bioassays predict the bioactive conformation of latent 143glycosyl‐IL‐6.

Low-affinity LFA1-dependent outside-in signaling mediates avidity modulation via the Rabin8-Rab8 axis.

Lymphocyte interactions mediated by leukocyte integrin lymphocyte function-associated antigen 1 (LFA1) and intercellular adhesion molecules (ICAMs) are important for lymphocyte trafficking and antigen recognition. Integrins are regulated by the modulation of ligand-binding affinity and avidity (valency). Although the mechanism underlying high-affinity LFA1 binding has been investigated extensively, the molecular mechanisms by which low-affinity multivalent binding initiates adhesion remain unclear. We previously showed that ICAM1 and monoclonal antibodies that recognize specific LFA1 conformations induce the accumulation of LFA1 at the contact surface. In this study, we found that the small GTPase Rab8 is critical for intracellular transport and accumulation of LFA1 at cell contact areas mediated by low-affinity LFA1-dependent outside-in signaling. Super-resolution microscopy revealed that Rab8 co-localized with LFA1 in small vesicles near the contact membrane. Inactivation of Rab8 decreased ICAM1-dependent adhesion and substantially reduced LFA1 density on the contact membrane. The GTP-bound active form of Rab8 increased cell adhesiveness and promoted LFA1 accumulation at the contact area through co-trafficking with LFA1. Rab8 activation was induced by low-affinity conformation-dependent outside-in signaling via the guanine exchange factor Rabin8, which induced Rab8 activation at the cell contact area independent of Rap1. Single-molecule imaging of ICAM1 on a supported planner lipid bilayer demonstrated that Rab8 increased the frequency of LFA1-ICAM1 interactions without affecting their binding lifetime, indicating that Rab8 is mainly involved in the modulation of LFA1 avidity rather than LFA1 affinity. The present findings underscore the importance of low-affinity conformation-dependent outside-in signaling via the Rabin8-Rab8 axis leading to the initiation of LFA1 transport to the contact area.

Abolished frameshifting for predicted structure-stabilizing SARS-CoV-2 mutants: implications to alternative conformations and their statistical structural analyses.

The SARS-CoV-2 frameshifting element (FSE) has been intensely studied and explored as a therapeutic target for coronavirus diseases, including COVID-19. Besides the intriguing virology, this small RNA is known to adopt many length-dependent conformations, as verified by multiple experimental and computational approaches. However, the role these alternative conformations play in the frameshifting mechanism and how to quantify this structural abundance has been an ongoing challenge. Here, we show by DMS and dual-luciferase functional assays that previously predicted FSE mutants (using the RAG graph theory approach) suppress structural transitions and abolish frameshifting. Furthermore, correlated mutation analysis of DMS data by three programs (DREEM, DRACO, and DANCE-MaP) reveals important differences in their estimation of specific RNA conformations, suggesting caution in the interpretation of such complex conformational landscapes. Overall, the abolished frameshifting in three different mutants confirms that all alternative conformations play a role in the pathways of ribosomal transition.

Interplay of PROTAC Complex Dynamics for Undruggable Targets: Insights into Ternary Complex Behavior and Linker Design.

Protein degraders, such as bifunctional proteolysis-targeting chimeras (PROTACs), selectively eliminate target proteins by leveraging the natural protein degradation machinery. PROTACs bridge the target protein with an E3 ligase, which induces ubiquitination and degradation. Investigating ternary complex structures elucidates the molecular mechanisms of their formation and degradation. This study examines the binding dynamics of E3 ligases (VHL, CRBN, and cIAP) with proteins of interest, focusing on dynamics, cooperativity, selectivity, linker length, and PROTAC conformations. The influence of interface residues and linker lengths on specific conformations for target proteins and E3 ligases is highlighted. Utilizing molecular dynamics and steered molecular dynamics simulations, the study provides comprehensive parameters on the behavior and stability of diverse ternary complexes. These insights are crucial for designing PROTACs targeting disease-causing proteins and advancing the development of degradable ternary complexes for therapeutic applications.

Collisional alignment and molecular rotation control the chemi-ionization of individual conformers of hydroquinone with metastable neon.

The relationship between the shape of a molecule and its chemical reactivity is a central tenet in chemistry. However, the influence of the molecular geometry on reactivity can be subtle and result from several opposing effects. Here, using a crossed-molecular-beam experiment in which individual rotational quantum states of specific conformers of a molecule are separated, we study the chemi-ionization reaction of hydroquinone with metastable neon atoms. We show that collision-induced alignment of the reaction partners caused by geometry-dependent long-range forces influences reaction pathways, which is, however, countered by molecular rotation. The present work provides insights into the conformation-specific stereodynamics of complex polyatomic systems and illustrates the capability of advanced molecule-control techniques to unravel these effects.

Advanced applications of Nanodiscs-based platforms for antibodies discovery

Due to their fundamental biological importance, membrane proteins (MPs) are attractive targets for drug discovery, with cell surface receptors, transporters, ion channels, and membrane-bound enzymes being of particular interest. However, due to numerous challenges, these proteins present underutilized opportunities for discovering biotherapeutics. Antibodies hold the promise of exquisite specificity and adaptability, making them the ideal candidates for targeting complex membrane proteins. They can target specific conformations of a particular membrane protein and can be engineered into various formats. Generating specific and effective antibodies targeting these proteins is no easy task due to several factors. The antigen's design, antibody-generation strategies, lead optimization technologies, and antibody modalities can be modified to tackle these challenges. The rational employment of cutting-edge lipid nanoparticle systems for retrieving the membrane antigen has been successfully implemented to simplify the mechanism-based therapeutic antibody discovery approach. Despite the highlighted MP production challenges, this review unequivocally underscores the advantages of targeting complex membrane proteins with antibodies and designing membrane protein antigens. Selected examples of lipid nanoparticle success have been illustrated, emphasizing the potential of therapeutic antibody discovery in this regard. With further research and development, we can overcome these challenges and unlock the full potential of therapeutic antibodies directed to target complex MPs.