👤 Sanjib Senapati

Cholesteryl ester transfer protein (CETP) is a crucial therapeutic target for combating cardiovascular disease (CVD) due to its strong influence in modulating high-density lipoprotein (HDL) levels. CETP is responsible for the bidirectional transfer of cholesteryl esters (CEs) and triglycerides (TGs) between different lipoprotein fractions. Although CETP encounters both these neutral lipid substrates when it penetrates deep into lipoprotein cores and can acquire either lipid, prior studies have examined its conformational space only in the presence of CEs or TGs individually. Here, we investigate the uncharacterised dynamics of CETP in heterogeneous lipid environments (CE-TG and TG-CE) using molecular dynamics simulations. Compared to the stable, homogeneous CE-bound state, the introduction of TG, particularly in mixed CE/TG configurations, induces significant structural instability and protein expansion. Mixed-lipid occupancy leads to elevated flexibility in critical lipoprotein-binding loops and the distortion of vital secondary structural elements. Furthermore, large-scale collective motion analyses reveal that heterogeneous binding forces CETP into aberrant, asymmetric, and hyper-twisted conformations. This disrupts the symmetric bending-twisting balance essential for efficient lipid exchange. Free energy landscapes confirm that the TGs within the mixed-lipid systems exhibit varied conformational states and adopt orientations that deviate from their reported parallel N-N orientation for lipid transfer through CETP. These findings suggest that the simultaneous presence of CE and TG imposes considerable conformational strain, fundamentally impairing CETP's lipid transport mechanism and offering novel mechanistic insights for future CETP-targeted therapeutics. Show less

Cholesteryl ester transfer protein (CETP) plays a central role in plasma lipid transport, facilitating the exchange of neutral lipids, such as cholesteryl esters and triglycerides, between lipoproteins. Despite the existence of several lipid-carrying/binding proteins in the family, such as lipopolysaccharide-binding protein (LBP), bacterial permeability increasing protein (BPI), and phospholipid transfer protein (PLTP), the structural and mechanistic uniqueness of CETP in neutral lipid transfer remains underexplored. Moreover, the involvement of PLTP in neutral lipid transfer is still debated, with researchers presenting conflicting mechanisms. Therefore, this study investigates the distinct structural ability of CETP in mediating neutral lipid exchange compared to other lipid-binding proteins. The study also emphasizes that simple protein modeling based on templates may not guarantee structural integrity unless validated through simulations. To achieve our objectives, we employed molecular docking, comparative molecular dynamics simulations, structural analysis, and lipid-protein interaction profiling with representative neutral lipids. In addition, protein-lipid affinities, tunnel architecture, and conformational flexibility were examined to characterize CETP's unique features and evaluate the quality of the constructed model for PLTP. The results demonstrated that a tunnel-like hydrophobic channel in CETP facilitates bidirectional neutral lipid transfer, unlike the compartmentalized binding pockets observed in other proteins. In addition, the neutral lipids' unfavorable conformational orientation was not affected in PLTP, whereas the same unfavorable conformation is changed to a favorable conformation in CETP, making only the lipid-carrying protein have the ability to transfer the neutral lipids. In conclusion, our findings highlight that the CETP is a specialized neutral lipid carrier with a unique structural mechanism distinct from typical lipid-binding proteins. This comparative insight enhances understanding of the structural plasticity of each lipid-carrying protein and the reliability of the modeled structure. Show less

Cardiovascular diseases (CVDs) have become a leading cause of deaths globally. Recent studies have shown that increasing the level of high-density lipoproteins (HDL) is one of the potential avenues to halt CVD progression. This could be achieved by modulating the neutral lipid transfer activity of cholesteryl ester transfer protein (CETP), a key target in developing effective cardioprotective drugs. This study aims to identify important structural fingerprints and functional moieties as "good" and "bad" contributors toward CETP inhibition, using machine learning (ML) and quantitative structure-activity relationship-based approaches. Results suggest unsaturated heterocyclic rings and trifluoromethyl substitutions as potential promoters and aliphatic carboxylic acid and ester moieties as the detractors in CETP inhibition. Molecular dynamics (MD) simulations of CETP in complexation with recently reported Obicetrapib with "good" fingerprints versus a clinically failed inhibitor, Torcetrapib shows superior inhibitory potential of the former due to stronger binding and better shape complementarity with the CETP hydrophobic tunnel. By leveraging the potentials of ML and MD simulations, this comprehensive study helps judicious pick of the right functional moieties for designing next generation CETP drugs targeting CVD. Show less

Human plasma cholesteryl ester transfer protein (CETP) mediates the transfer of neutral lipids from antiatherogenic high-density lipoproteins (HDLs) to proatherogenic low-density lipoproteins (LDLs). Recent cryo-electron microscopy studies have suggested that CETP penetrates its N- and C-terminal domains in HDL and LDL to form a ternary complex, which facilitates the lipid transfer between different lipoproteins. Inhibition of CETP lipid transfer activity has been shown to increase the plasma HDL-C levels and, therefore, became an effective strategy for combating cardiovascular diseases. Thus, understanding the molecular mechanism of inhibition of lipid transfer through CETP is of paramount importance. Recently reported inhibitors, torcetrapib and anacetrapib, exhibited low potency in addition to severe side effects, which essentially demanded a thorough knowledge of the inhibition mechanism. Here, we employ steered molecular dynamics simulations to understand how inhibitors interfere with the neutral lipid transfer mechanism of CETP. Our study revealed that inhibitors physically occlude the tunnel posing a high energy barrier for lipid transfer. In addition, inhibitors bring about the conformational changes in CETP that hamper CE passage and expose protein residues that disrupt the optimal hydrophobicity of the CE transfer path. The atomic level details presented here could accelerate the designing of safe and efficacious CETP inhibitors. Show less

Cholesteryl ester transfer protein (CETP) facilitates the transfer of cholesteryl esters (CEs) from antiatherogenic high-density lipoproteins to proatherogenic low-density lipoproteins. Inhibition of CETP is therefore being pursued as a potential strategy to reduce cardiovascular risk. The crystal structure of CETP has revealed the existence of two neutral CEs and two charged phospholipids (PLs) in its hydrophobic tunnel. This is in direct contrast to the other lipid-binding proteins that contain only two bound lipids. Moreover, previous animal studies on mice showed no detectable PL-transfer activity of CETP. Thus, the role of bound PLs in CETP is completely unknown. Here, we employ molecular dynamics simulations and free-energy calculations to unravel the primary effects of bound PLs on CETP structure and dynamics and attempt to correlate the observed changes to its function. Our results suggest that the structure of CETP is elastic and can attain different conformations depending on the state of bound PLs. In solution, these PLs maintain CETP in a bent-untwisted conformation that can uphold neutral lipids in its core tunnel. Results also suggest that although both PLs complement each other in their action, the C-terminal PL (C-PL) imparts greater influence on CETP by virtue of its tighter binding. Our finding fits very well with the recent inhibitor-bound CETP crystal structure, where the inhibitor displaced the N-terminal PL for binding to CETP's central domain without disrupting the binding of C-PL. We speculate that the observed increased flexibility of CETP in the absence of PLs could play a crucial role in its binding with lipoproteins and subsequent lipid-transfer activity. Show less

CETP transfers cholesteryl esters (CEs) and triglycerides (TGs) between different lipoproteins and came in limelight as a drug-target against CVD. In the search for detailed mechanism of lipid transfer through CETP, enormous effort is devoted employing crystallographic, cryo-EM, and Molecular Dynamics (MD) studies. However, these studies primarily focused on CE-bound CETP structure and CE transfer mechanism. With the reported correlation that CETP looses significant CE transfer activity upon inhibiting TG transfer, it is of tremendous importance to understand the structure and dynamics of TG-bound CETP. Our results from large-scale all-atom and coarse-grained MD simulations show that CETP can accommodate two TG molecules in parallel N-N orientation with TG oleate chains majorly attaining the tuning-fork conformation. In TG-bound form, CETP not only maintained its secondary structures but also exhibited similar bending-twisting motions as reported for CE-CETP crystal structure. Obtained structural information are further validated by correlating to available functional data of 2-8 fold slower transfer rate of TG through CETP, where we show that TGs make 20% additional contacts with CETP compared to CEs. Identified CETP residues facilitating TG binding also match very well with reported mutagenesis data. The study could accelerate the drug-designing processes to combat CETP functionality and CVD. Show less