Crystal structure reveals the hydrophilic R1 group impairs NDM-1–ligand binding via water penetration at L3
The global spread of New Delhi metallo-β-lactamases (NDMs) has exacerbated the antimicrobial resistance crisis. This study resolved the crystal structure of NDM-1 hydrolyzing amoxicillin for the first time, revealed that the hydroxyl group in the R1 moiety of amoxicillin anchors a key water molecule...
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Elsevier
2025-12-01
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| Series: | Journal of Structural Biology: X |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2590152425000145 |
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| author | Xiangrui Shi Huijuan Yang Yujie Dai Hui Zhao Yuhang Li Yanxi Li Xin Zhou Hailong Yan Qinghua Zhang Wei Liu |
| author_facet | Xiangrui Shi Huijuan Yang Yujie Dai Hui Zhao Yuhang Li Yanxi Li Xin Zhou Hailong Yan Qinghua Zhang Wei Liu |
| author_sort | Xiangrui Shi |
| collection | DOAJ |
| description | The global spread of New Delhi metallo-β-lactamases (NDMs) has exacerbated the antimicrobial resistance crisis. This study resolved the crystal structure of NDM-1 hydrolyzing amoxicillin for the first time, revealed that the hydroxyl group in the R1 moiety of amoxicillin anchors a key water molecule (Wat1) via hydrogen bond, inducing a conformational shift in Met67 (average displacement of 3.8 Å compared to its position in complexes with ampicillin, penicillin G, and penicillin V) and impairing the hydrophobic interaction between the loop 3 and the substrate. Molecular dynamics simulations confirmed that the π-π stacking contact time between amoxicillin and the L3 critical residue Phe70 decreased to 4.3 % (ampicillin: 12.3 %), with a binding energy reduction of 10.5 kcal/mol. Steady-state kinetics showed that amoxicillin exhibited a 2.2-fold higher Km and a 5.2-fold higher kcat compared to ampicillin, demonstrating that hydrophilic R1 groups impair enzyme-substrate binding. This work demonstrates the essential role of hydrophobic interactions in L3-mediated substrate binding and provides a novel strategy for designing L3-targeted NDM-1 inhibitors: maximize hydrophobicity and minimize polar surface area in the L3 contact region to block water penetration, thereby stabilizing the inhibitor-L3 interaction. |
| format | Article |
| id | doaj-art-3308d59dc0414bc89eda1504c59b2c69 |
| institution | Kabale University |
| issn | 2590-1524 |
| language | English |
| publishDate | 2025-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Structural Biology: X |
| spelling | doaj-art-3308d59dc0414bc89eda1504c59b2c692025-08-20T03:31:06ZengElsevierJournal of Structural Biology: X2590-15242025-12-011210013310.1016/j.yjsbx.2025.100133Crystal structure reveals the hydrophilic R1 group impairs NDM-1–ligand binding via water penetration at L3Xiangrui Shi0Huijuan Yang1Yujie Dai2Hui Zhao3Yuhang Li4Yanxi Li5Xin Zhou6Hailong Yan7Qinghua Zhang8Wei Liu9Chongqing Key Laboratory of Natural Product Synthesis and Drug Research, School of Pharmaceutical Sciences, Chongqing University, Chongqing 401331, ChinaChongqing Key Laboratory of Natural Product Synthesis and Drug Research, School of Pharmaceutical Sciences, Chongqing University, Chongqing 401331, ChinaDepartment of Obstetrics and Gynecology, Daping Hospital, Army Medical University, Chongqing 400042, ChinaNational Engineering Research Center of Immunological Products, Department of Microbiology and Biochemical Pharmacy, College of Pharmacy, Army Medical University, Chongqing 400038, ChinaNational Engineering Research Center of Immunological Products, Department of Microbiology and Biochemical Pharmacy, College of Pharmacy, Army Medical University, Chongqing 400038, ChinaDepartment of Obstetrics and Gynecology, Daping Hospital, Army Medical University, Chongqing 400042, ChinaDepartment of Obstetrics and Gynecology, Daping Hospital, Army Medical University, Chongqing 400042, ChinaChongqing Key Laboratory of Natural Product Synthesis and Drug Research, School of Pharmaceutical Sciences, Chongqing University, Chongqing 401331, China; Corresponding authors.Department of Obstetrics and Gynecology, Daping Hospital, Army Medical University, Chongqing 400042, China; Corresponding authors.Department of Obstetrics and Gynecology, Daping Hospital, Army Medical University, Chongqing 400042, China; Institute of Immunology, Army Medical University, Chongqing 400038, China; Corresponding author at: Department of Obstetrics and Gynecology, Daping Hospital, Army Medical University, Chongqing 400042, China.The global spread of New Delhi metallo-β-lactamases (NDMs) has exacerbated the antimicrobial resistance crisis. This study resolved the crystal structure of NDM-1 hydrolyzing amoxicillin for the first time, revealed that the hydroxyl group in the R1 moiety of amoxicillin anchors a key water molecule (Wat1) via hydrogen bond, inducing a conformational shift in Met67 (average displacement of 3.8 Å compared to its position in complexes with ampicillin, penicillin G, and penicillin V) and impairing the hydrophobic interaction between the loop 3 and the substrate. Molecular dynamics simulations confirmed that the π-π stacking contact time between amoxicillin and the L3 critical residue Phe70 decreased to 4.3 % (ampicillin: 12.3 %), with a binding energy reduction of 10.5 kcal/mol. Steady-state kinetics showed that amoxicillin exhibited a 2.2-fold higher Km and a 5.2-fold higher kcat compared to ampicillin, demonstrating that hydrophilic R1 groups impair enzyme-substrate binding. This work demonstrates the essential role of hydrophobic interactions in L3-mediated substrate binding and provides a novel strategy for designing L3-targeted NDM-1 inhibitors: maximize hydrophobicity and minimize polar surface area in the L3 contact region to block water penetration, thereby stabilizing the inhibitor-L3 interaction.http://www.sciencedirect.com/science/article/pii/S2590152425000145NDM-1AmoxicillinL3R1 group |
| spellingShingle | Xiangrui Shi Huijuan Yang Yujie Dai Hui Zhao Yuhang Li Yanxi Li Xin Zhou Hailong Yan Qinghua Zhang Wei Liu Crystal structure reveals the hydrophilic R1 group impairs NDM-1–ligand binding via water penetration at L3 Journal of Structural Biology: X NDM-1 Amoxicillin L3 R1 group |
| title | Crystal structure reveals the hydrophilic R1 group impairs NDM-1–ligand binding via water penetration at L3 |
| title_full | Crystal structure reveals the hydrophilic R1 group impairs NDM-1–ligand binding via water penetration at L3 |
| title_fullStr | Crystal structure reveals the hydrophilic R1 group impairs NDM-1–ligand binding via water penetration at L3 |
| title_full_unstemmed | Crystal structure reveals the hydrophilic R1 group impairs NDM-1–ligand binding via water penetration at L3 |
| title_short | Crystal structure reveals the hydrophilic R1 group impairs NDM-1–ligand binding via water penetration at L3 |
| title_sort | crystal structure reveals the hydrophilic r1 group impairs ndm 1 ligand binding via water penetration at l3 |
| topic | NDM-1 Amoxicillin L3 R1 group |
| url | http://www.sciencedirect.com/science/article/pii/S2590152425000145 |
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