Structural basis for selective inhibition of Src family kinases by PP1.

BACKGROUND: Small-molecule inhibitors that can target individual kinases are powerful tools for use in signal transduction research. It is difficult to find such compounds because of the enormous number of protein kinases and the highly conserved nature of their catalytic domains. Recently, a novel, potent, Src family selective tyrosine kinase inhibitor was reported (PP1). Here, we study the structural basis for this inhibitor's specificity for Src family kinases. RESULTS: A single residue corresponding to Ile338 (v-Src numbering; Thr338 in c-Src) in Src family tyrosine kinases largely controls PP1's ability to inhibit protein kinases. Mutation of Ile338 to a larger residue such as methionine or phenylalanine in v-Src makes this inhibitor less potent. Conversely, mutation of Ile338 to alanine or glycine increases PP1's potency. PP1 can inhibit Ser/Thr kinases if the residue corresponding to Ile338 in v-Src is mutated to glycine. We have accurately predicted several non-Src family kinases that are moderately (IC(50) approximately 1 microM) inhibited by PP1, including c-Abl and the MAP kinase p38. CONCLUSIONS: Our mutagenesis studies of the ATP-binding site in both tyrosine kinases and Ser/Thr kinases explain why PP1 is a specific inhibitor of Src family tyrosine kinases. Determination of the structural basis of inhibitor specificity will aid in the design of more potent and more selective protein kinase inhibitors. The ability to desensitize a particular kinase to PP1 inhibition of residue 338 or conversely to sensitize a kinase to PP1 inhibition by mutation should provide a useful basis for chemical genetic studies of kinase signal transduction.     

Reversible Inhibitors Arrest ClpP in a Defined Conformational State that Can Be Revoked by ClpX Association

Caseinolytic protease P (ClpP) is an important regulator of Staphylococcus aureus pathogenesis. A high‐throughput screening for inhibitors of ClpP peptidase activity led to the identification of the first non‐covalent binder for this enzyme class. Co‐crystallization of the small molecule with S. aureus ClpP revealed a novel binding mode: Because of the rotation of the conserved residue proline 125, ClpP is locked in a defined conformational state, which results in distortion of the catalytic triad and inhibition of the peptidase activity. Based on these structural insights, the molecule was optimized by rational design and virtual screening, resulting in derivatives exceeding the potency of previous ClpP inhibitors. Strikingly, the conformational lock is overturned by binding of ClpX, an associated chaperone that enables proteolysis by substrate unfolding in the ClpXP complex. Thus, regulation of inhibitor binding by associated chaperones is an unexpected mechanism important for ClpP drug development.     

Structural insight into exosite binding and discovery of novel exosite inhibitors of botulinum neurotoxin serotype A through in silico screening

Botulinum neurotoxin serotype A (BoNT/A) is the most lethal toxin among the Tier 1 Select Agents. Development of potent and selective small molecule inhibitors against BoNT/A zinc metalloprotease remains a challenging problem due to its exceptionally large substrate binding surface and conformational plasticity. The exosites of the catalytic domain of BoNT/A are intriguing alternative sites for small molecule intervention, but their suitability for inhibitor design remains largely unexplored. In this study, we employed two recently identified exosite inhibitors, D-chicoric acid and lomofungin, to probe the structural features of the exosites and molecular mechanisms of synergistic inhibition. The results showed that D-chicoric acid favors binding at the α-exosite, whereas lomofungin preferentially binds at the β-exosite by mimicking the substrate β-sheet binding interaction. Molecular dynamics simulations and binding interaction analysis of the exosite inhibitors with BoNT/A revealed key elements and hotspots that likely contribute to the inhibitor binding and synergistic inhibition. Finally, we performed database virtual screening for novel inhibitors of BoNT/A targeting the exosites. Hits C1 and C2 showed non-competitive inhibition and likely target the α- and β-exosites, respectively. The identified exosite inhibitors may provide novel candidates for structure-based development of therapeutics against BoNT/A intoxication.     

A Src SH2 selective binding compound inhibits osteoclast-mediated resorption

BACKGROUND: The observations that Src(-/-) mice develop osteopetrosis and Src family tyrosine kinase inhibitors decrease osteoclast-mediated resorption of bone have implicated Src in the regulation of osteoclast-resorptive activity. We have designed and synthesized a compound, AP22161, that binds selectively to the Src SH2 domain and demonstrated that it inhibits Src-dependent cellular activity and inhibits osteoclast-mediated resorption. RESULTS: AP22161 was designed to bind selectively to the Src SH2 domain by targeting a cysteine residue within the highly conserved phosphotyrosine-binding pocket. AP22161 was tested in vitro for binding to SH2 domains and was found to bind selectively and with high affinity to the Src SH2 domain. AP22161 was further tested in mechanism-based cellular assays and found to block Src SH2 binding to peptide ligands, inhibit Src-dependent cellular activity and diminish osteoclast resorptive activity. CONCLUSIONS: These results indicate that a compound that selectively inhibits Src SH2 binding can be used to inhibit osteoclast resorption. Furthermore, AP22161 has the potential to be further developed for treating osteoporosis.     

多肽抑制剂抑制淀粉质多肽42构象转换的分子动力学模拟和结合自由能计算

应用分子动力学模拟和结合自由能计算方法研究了多肽抑制剂KLVFF、VVIA和LPFFD抑制淀粉质多肽42 (Aβ42)构象转换的分子机理. 结果表明, 三种多肽抑制剂均能够有效抑制Aβ42的二级结构由α-螺旋向β-折叠的构象转换. 另外, 多肽抑制剂降低了Aβ42分子内的疏水相互作用, 减少了多肽分子内远距离的接触, 有效抑制了Aβ42的疏水塌缩, 从而起到稳定其初始构象的作用. 这些抑制剂与Aβ42之间的疏水和静电相互作用(包括氢键)均有利于它们抑制Aβ42的构象转换. 此外, 抑制剂中的带电氨基酸残基可以增强其和Aβ42之间的静电相互作用(包括氢键), 并降低抑制剂之间的聚集, 从而大大增强对Aβ42构象转换的抑制能力. 但脯氨酸的引入会破坏多肽的线性结构, 从而大大降低其与Aβ42 之间的作用力. 上述分子模拟的结果揭示了多肽抑制剂KLVFF、VVIA和LPFFD抑制Aβ42构象转换的分子机理, 对于进一步合理设计Aβ的高效短肽抑制剂具有非常重要的理论指导意义.     

Calorimetric and structural studies of 1,2,3-trisubstituted cyclopropanes as conformationally constrained peptide inhibitors of Src SH2 domain binding.

Isothermal titration calorimetry and X-ray crystallography have been used to determine the structural and thermodynamic consequences associated with constraining the pTyr residue of the pYEEI ligand for the Src Homology 2 domain of the Src kinase (Src SH2 domain). The conformationally constrained peptide mimics that were used are cyclopropane-derived isosteres whereby a cyclopropane ring substitutes to the N-Calpha-Cbeta atoms of the phosphotyrosine. Comparison of the thermodynamic data for the binding of the conformationally constrained peptide mimics relative to their equivalent flexible analogues as well as a native tetrapeptide revealed an entropic advantage of 5-9 cal mol(-1) K(-1) for the binding of the conformationally constrained ligands. However, an unexpected drop in enthalpy for the binding of the conformationally constrained ligands relative to their flexible analogues was also observed. To evaluate whether these differences reflected conformational variations in peptide binding modes, we have determined the crystal structure of a complex of the Src SH2 domain bound to one of the conformationally constrained peptide mimics. Comparison of this new structure with that of the Src SH2 domain bound to a natural 11-mer peptide (Waksman et al. Cell 1993, 72, 779-790) revealed only very small differences. Hence, cyclopropane-derived peptides are excellent mimics of the bound state of their flexible analogues. However, a rigorous analysis of the structures and of the surface areas at the binding interface, and subsequent computational derivation of the energetic binding parameters, failed to predict the observed differences between the binding thermodynamics of the rigidified and flexible ligands, suggesting that the drop in enthalpy observed with the conformationally constrained peptide mimic arises from sources other than changes in buried surface areas, though the exact origin of the differences remains unclear.     

Structure-based design and evaluation of novel N-phenyl-1H-indol-2-amine derivatives for fat mass and obesity-associated (FTO) protein inhibition

Fat mass and obesity-associated (FTO) protein contributes to non-syndromic human obesity which refers to excessive fat accumulation in human body and results in health risk. FTO protein has become a promising target for anti-obesity medicines as there is an immense need for the rational design of potent inhibitors to treat obesity. In our study, a new scaffold N-phenyl-1H-indol-2-amine was selected as a base for FTO protein inhibitors by applying scaffold hopping approach. Using this novel scaffold, different derivatives were designed by extending scaffold structure with potential functional groups. Molecular docking simulations were carried out by using two different docking algorithm implemented in CDOCKER (flexible docking) and AutoDock programs (rigid docking). Analyzing results of rigid and flexible docking, compound MU06 was selected based on different properties and predicted binding affinities for further analysis. Molecular dynamics simulation of FTO/MU06 complex was performed to characterize structure rationale and binding stability. Certainly, Arg96 and His231 residue of FTO protein showed stable interaction with inhibitor MU06 throughout the production dynamics phase. Three residues of FTO protein (Arg96, Asp233, and His231) were found common in making H-bond interactions with MU06 during molecular dynamics simulation and CDOCKER docking.     

Structural and energetic insight into the interactions between the benzolactam inhibitors and tumor marker HSP90α

The heat shock protein 90α (HSP90α) provides a promising molecular target for cancer therapy. A series of novel benzolactam inhibitors exhibited distinct inhibitory activity for HSP90α. However, the structural basis for the impact of distinct R₁ substituent groups of nine benzolactam inhibitors on HSP90α binding affinities remains unknown. In this study, we carried out molecular docking, molecular dynamics (MD) simulations, and molecular mechanics and generalized Born/surface area (MM–GBSA) binding free energy calculations to address the differences. Molecular docking studies indicated that all nine compounds presented one conformation in the ATP-binding site of HSP90α N-terminal domain. MD simulations and subsequent MM–GBSA calculations revealed that the hydrophobic interactions between all compounds and HSP90α contributed the most to the binding affinity and a good linear correlation was obtained between the calculated and the experimental binding free energies (R = 0.88). The per residue decomposition revealed that the most remarkable differences of residue contributions were found in the residues Ala55, Ile96, and Leu107 defining a hydrophobic pocket for the R₁ group, consistent with the analysis of binding modes. This study may be helpful for the future design of novel HSP90α inhibitors.     

Structure-based design of Aurora A & B inhibitors

The Aurora family of serine/threonine kinases are mitotic regulators involved in centrosome duplication, formation of the bipolar mitotic spindle and the alignment of the chromosomes along the spindle. These proteins are frequently overexpressed in tumor cells as compared to normal cells and are therefore potential therapeutic oncology targets. An Aurora A high throughput screen revealed a promising sub-micromolar indazole-benzimidazole lead. Modification of the benzimidazole portion of the lead to a C2 linker with a phenyl ring was proposed to achieve novelty. Docking revealed that a conjugated linker was optimal and the resulting compounds were equipotent with the lead. Further structure-guided optimization of substituents on the 5 & 6 position of the indazole led to single digit nanomolar potency. The homology between the Aurora A & Aurora B kinase domains is 71% but their binding sites only differ at residues 212 & 217 (Aurora A numbering). However interactions with only the latter residue may be used for obtaining selectivity. An analysis of published Aurora A and Aurora B X-ray structures reveals subtle differences in the shape of the binding sites. This was exploited by introduction of appropriately sized substituents in the 4 & 6 position of the indazole leading to Aurora B selective inhibitors. Finally we calculate the conformational energy penalty of the putative bioactive conformation of our inhibitors and show that this property correlates well with the Aurora A binding affinity.     

Conformational changes in the metallo-beta-lactamase ImiS during the catalytic reaction: an EPR spectrokinetic study of Co(II)-spin label interactions

Metallo-beta-lactamases are responsible for conferring antibiotic resistance on certain pathogenic bacteria. In consequence, the search for inhibitors that may be useful in combating antibiotic resistance has fueled much study of the active sites of these enzymes. There exists circumstantial evidence that the binding of substrates and inhibitors to metallo-beta-lactamases may involve binding to the organic part of the molecule, in addition to or prior to binding to one or more active site metal ions. It has also been postulated that a conformational change may accompany this putative binding. In the present study, electron paramagnetic resonance spectrokinetic study of a spin-labeled variant of the class B2 metallo-beta-lactamase ImiS identified movement of a component residue on a conserved alpha-helix in a catalytically competent time upon formation of a transient reaction intermediate with the substrate imipenem. In a significant subpopulation of ImiS, this conformational change was not associated with substrate binding to the active site metal ion but, rather, represents a distinct step in the reaction with ImiS. This observation has implications regarding the determinants of substrate specificity in metallo-beta-lactamases and the design of potentially clinically useful inhibitors.     

Tailored Peptide Phenyl Esters Block ClpXP Proteolysis by an Unusual Breakdown into a Heptamer–Hexamer Assembly

The proteolytic complex ClpXP is fundamental to bacterial homeostasis and pathogenesis. Because of its conformational flexibility, the development of potent ClpXP inhibitors is challenging, and novel tools to decipher its intricate regulation are urgently needed. Herein, we present amino acid based phenyl esters as molecular probes to study the activity and oligomerization of the ClpXP complex of S. aureus. Systematic screening of (R)‐ and (S)‐amino acids led to compounds showing potent inhibition, as well as stimulation of ClpXP‐mediated proteolysis. Substoichiometric binding of probes arrested ClpXP in an unprecedented heptamer–hexamer assembly, in which the two heptameric ClpP rings are dissociated from each other. At the same time, the affinity between ClpX and ClpP increased, leading to inhibition of both enzymes. This conformational arrest is beneficial for the consolidated shutdown of ClpXP, as well as for the study of the oligomeric state during its catalytic cycle.     

Cyclopropane-derived peptidomimetics. design, synthesis, and evaluation of novel Ras farnesyltransferase inhibitors

Trisubstituted cyclopropanes have previously been established as rigid replacements of dipeptide arrays in several biological systems. Toward further evaluating the utility of these dipeptide mimics in the design of novel CA(1)A(2)X-based inhibitors of Ras farnesyltransferase (FTase), the conformationally constrained, diastereomeric pseudopeptides CAbuPsi[COcpCO]FM 7-9, the flexible analogue CAbuPsi[CHOHCH(2)]FM (10), and the tetrapeptide CAbuFM (6) were prepared. The orientations of the two peptide backbone substituents and the phenyl group on the cyclopropane rings in 7-9were specifically designed to probe selected topological features of the hydrophobic binding pocket of the A(2) subsite of FTase. The syntheses of the requisite trisubstituted cyclopropane carboxylic acid 22 and the diastereomeric cyclopropyl lactones 32a,b featured diastereoselective intramolecular cyclopropanations of chiral allylic diazoacetates and a new method for introducing side chains onto the C-terminal amino acid of cyclopropane-derived dipeptide replacements via the opening of an N-Boc-aziridine with an organocuprate. These cyclopropane intermediates were then converted into the targeted FTase inhibitors 7-9 by standard peptide coupling techniques. The pseudopeptides 7-9 were found to be competitive inhibitors of Ras FTase with IC(50)s of 1055 nM for 7, 760 nM for 8, and 7200 nM for 9. The flexible analogue 10 of these constrained inhibitors exhibited a IC(50) of 320 nM and hence was slightly more potent than 7 and 8. All of these pseudopeptides were less potent than the tetrapeptide parent CAbuFM (6), which had an IC(50) of 38 nM. Because 7 and 8 are approximately equipotent, it appears that the orientation of the peptide backbone substituents on the cyclopropane rings in 7 and 8 do not have any significant effect on binding affinity and that multiple binding modes are possible without significant changes in affinity. On the other hand, this flexibility does not extend to the orientation of the side chain of the A(2) residue as 7 and 8 were both nearly 1 order of magnitude more potent than 9. Comparison of the relative potencies of 6 and 10 suggests that the amide linkage between the A(1) and the A(2) residues of CA(1)A(2)X-derived FTase inhibitors is important.     

Theoretical studies on the conformational change of adenosine kinase induced by inhibitors

Adenosine kinase (AK) is a two‐domain protein that catalyzes the phosphorylation of adenosine to adenosine monophosphate. Inhibitors of AK could increase adenosine to levels that activate nearby adenosine receptors and produce a wide variety of therapeutically beneficial activities. To get insight into the interaction mechanism between inhibitors and AK, we chose two kinds of novel inhibitors, alkynylpyrimidine inhibitor (APy) and aryl‐nucleoside inhibitor (AN), and used docking and molecular dynamics simulation methods to study the conformational changes of human AK on binding inhibitors. The calculation results revealed that both APy and AN could induce conformational changes of AK and stabilize AK at different semiopen conformations. On binding APy, the small lid‐domain rotated 14°, and the binding pocket rearranged after MD simulation. But in AK‐AN complex, the rotation of small domain is 22°, and the sugar ring of AN is mobile in the binding pocket. Further docking calculations on APy analogues indicate that the semiopen conformation could well explain the SAR of AK inhibitors. © 2010 Wiley Periodicals, Inc. Int J Quantum Chem, 2011     

Novel Dual‐Site‐Binding Neuraminidase Inhibitor from Virtual Screening by Pharmacophore and Molecular Dynamics Methods

Neuraminidase is a significant anti‐influenza target that plays crucial role in virus replication cycle. The discovery of 150‐cavity in Group‐1 neuraminidase provides us a novel mentality of designing inhibitor which can bind with both conserved site and 150‐cavity. In order to discover novel dual‐site‐binding inhibitors, a 3D chemical‐feature‐based pharmacophore model was established to cover dual‐site in neuraminidase. The dual‐site‐binding model was consistent in predicting the binding conformation of Group‐1 neuraminidase inhibitor and applied for virtual screening of Specs database. Compound 4 (ZINC05790048) that aligned well to the model was selected after multiple filtrations for molecular dynamics simulations, indicating improved binding energy with neuraminidase. It can sever as the lead compound for a novel series of inhibitors.     

Y220C突变体影响p53C蛋白质构象转换的分子动力学模拟

p53是迄今发现突变频率最高的一种肿瘤抑制蛋白质,突变会导致p53抑癌功能丧失并诱导癌症的发生。绝大多数的突变发生在p53的核心DNA结合区域（p53C）,其中Y220C是研究较多的一种突变体。虽然已有研究表明该突变能够降低p53C的结构稳定性,但其影响p53C构象转换的分子机制尚不清晰。本文利用分子动力学（MD）模拟方法研究了p53C突变体Y220C（p53C-Y220C）的结构变化,发现Y220C突变主要影响Y220C cluster区域（包括残基138-164和215-238）,且Y220C突变减少了Y220C cluster的β-折叠含量。进一步分析发现,Y220C突变不仅直接破坏突变氨基酸与周围氨基酸Leu145和Thr155之间的氢键,而且降低了Y220C cluster区域的折叠片S3和S8之间的氢键数量,使Y220C突变所形成的亲水性空腔变大,加速了水分子进入该蛋白质内部,并最终导致了p53C-Y220C变性。MD模拟结果揭示了Y220C突变影响p53C结构转换的分子机制,该研究对p53C-Y220C突变体高效稳定剂的筛选和设计具有重要意义。     

Dissecting the molecular recognition of dual lapatinib derivatives for EGFR/HER2

Abnormalities in the expression levels of EGFR/HER2 are found in many different types of human cancer; therefore, the design of dual inhibitors of EGFR/HER2 is a recognized anti-cancer strategy. Some lapatinib derivatives have been previously synthesized by modification at the methylsulfonylethylaminomethylfuryl group and biologically evaluated, demonstrating that the 2i compound shows potent inhibitory activity against EGFR/HER2-overexpressing cancer cells. In the present study, we explored the structural and energetic features that guide the molecular recognition of 2i using various EGFR/HER2 states. Molecular dynamics (MD) simulation with an MMPB(GB)SA approach was used to generate the inactive EGFR/HER2–ligand complexes. Our results corroborate that slight modification of lapatinib contributes to an increase in the affinity of the 2i compound for inactive EGFR/HER2 as compared with lapatinib compound, which is in accordance with experimental results. Comparison with previous results reveals that lapatinib and its derivative bind more strongly to the inactive than the intermediate active-inactive HER2 state. Principal component analysis allowed the observation that coupling of 2i to EGFR/HER2 is linked to a reduction in the conformational mobility, which may also contribute to the improvement in affinity observed for this compound as compared with lapatinib.     

Targeting the conformational transitions of MDM2 and MDMX: insights into dissimilarities and similarities of p53 recognition

MDM2 and MDMX are oncogenic homologue proteins that regulate the activity and stability of p53, a tumor suppressor protein involved in more than 50% of human cancers. While the large body of experiments so far accumulated has validated MDM2 as a therapeutically important target for the development of anticancer drugs, it is only recently that MDMX has also become an attractive target for the treatment of tumor cells expressing wild type p53. The availability of structural information of the N-terminal domain of MDM2 in complex with p53-derived peptides and inhibitors, and the very recent disclosure of the crystal structure of the N-terminal domain of MDMX bound to a p53 peptide, offer an unprecedented opportunity to provide insight into the molecular basis of p53 recognition and the identification of discriminating features affecting the binding of the tumor suppressor protein at MDM2 and MDMX. By using coarse graining simulations, in this study we report the exploration of the conformational transitions featured in the pathway leading from the apo-MDM2 and apo-MDMX states to the p53-bound MDM2 and p53-bound MDMX states, respectively. The results have enabled us to identify a pool of diverse conformational states of the oncogenic proteins that affect the binding of p53 and the presence of conserved and non-conserved interactions along the conformational transition pathway that may be exploited in the design of selective and dual modulators of MDM2 and MDMX activity.     

Mutation effects of neuraminidases and their docking with ligands: a molecular dynamics and free energy calculation study

A systematic study has been performed on neuraminidase (NA) mutations and NA-inhibitor docked complexes, with the aim to understand protein–ligand interactions and design broad-spectrum antiviral drugs with minimal resistances. The catalytic D151 residue is likely to mutate while others are relatively conserved. The NA active-site conformations are altered by mutations, but more alterations do not necessarily result in larger deviations to the binding properties. The effects of all related mutations have been discussed; e.g., for the arginine triad (R118, R292 and R371), it is found that residue R118 plays the most significant role during ligand binding. Generally, the calculated binding free energies agree well with the experimental observations. Susceptibility of influenza virus to NA inhibitors can be reinforced by some mutations; e.g., the binding free energies of ligands with N2 subtype increase from ?18.0 to ?42.1 kcal mol^?1 by the E119D mutation. Mutations of the various NA subtypes often cause similar conformational and binding changes, explaining the occurrence of cross resistances; nonetheless, differences can be detected in some cases that correspond to subtype-specific resistances. For all NA subtypes, the electrostatic contributions are the major driving force for ligand binding and largely responsible for the binding differences between the wild-type and mutated NA proteins.     

Connectivity and binding‐site recognition: Applications relevant to drug design

Here, we describe a family of methods based on residue–residue connectivity for characterizing binding sites and apply variants of the method to various types of protein–ligand complexes including proteases, allosteric‐binding sites, correctly and incorrectly docked poses, and inhibitors of protein–protein interactions. Residues within ligand‐binding sites have about 25% more contact neighbors than surface residues in general; high‐connectivity residues are found in contact with the ligand in 84% of all complexes studied. In addition, a k‐means algorithm was developed that may be useful for identifying potential binding sites with no obvious geometric or connectivity features. The analysis was primarily carried out on 61 protein–ligand structures from the MEROPS protease database, 250 protein–ligand structures from the PDBSelect (25%), and 30 protein–protein complexes. Analysis of four proteases with crystal structures for multiple bound ligands has shown that residues with high connectivity tend to have less variable side‐chain conformation. The relevance to drug design is discussed in terms of identifying allosteric‐binding sites, distinguishing between alternative docked poses and designing protein interface inhibitors. Taken together, this data indicate that residue–residue connectivity is highly relevant to medicinal chemistry. © 2010 Wiley Periodicals, Inc. J Comput Chem, 2010     

VEGFR2活性腔性质以及与抑制剂的结合模式研究

VEGFR2介导肿瘤诱导的血管生成作用, 是抑制肿瘤生长和转移的新靶点. 为深入探讨VEGFR2活性腔性质以及与抑制剂的结合模式, 采用多拷贝同时搜寻法(MCSS)研究VEGFR2活性腔的性质, 然后用分子对接方法对5个已上临床的VEGFR抑制剂与VEGFR2活性腔进行对接计算, 讨论它们的结合模式, 确定与配体结合相关的关键残基. 研究发现: 疏水腔I, II是配体结合的关键区域, 残基Glu915, Cys917是关键的氢键作用位点, Lys866, Glu883和Asp1044形成的极性区域对提高配体亲合力很重要, 疏水腔III和极性腔IV是额外增强配体结合力的区域, IV区的Arg1030可提供额外的氢键作用位点. 本研究可为全新VEGFR2抑制剂的合理药物设计提供理论依据, 为寻找新的抗肿瘤药物奠定基础.