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Different binding modes of stereoisomers of beta-lactamase inhibitors by flexible docking

Title
Different binding modes of stereoisomers of beta-lactamase inhibitors by flexible docking
Authors
목성희
Issue Date
2006
Department/Major
대학원 약학과
Publisher
이화여자대학교 대학원
Degree
Master
Abstract
The β-lactamases confer antibiotic resistance to bacteria by catalyzing the hydrolysis of β-lactam antibiotics. β-lactamase inhibitors are therefore of considerable interest in the battle against bacteria. β-lactamases are classified into two major groups on the basis of their catalytic mechanism. The first group that requires serine in the active site is consisted of classes A and C, and the second group that requires metal, class B. Class C and class B β-lactamases are among the most problematic of these enzymes. Not only are they widely expressed among nosocomial pathogens but also they are not significantly inhibited by clinically used β-lactamase inhibitors. Using the flexible docking program, FlexiDock and QXP, two sets of α and β isomers of novel allyl substituted derivatives of sulbactam were docked into the Bacteroides fragilis metallo-β-lactamase and four diastereomers of 3-Phenyl-clavams, four diastereomers of 6-Methyl 3-Phenyl clavams and two sets of D and L novel phosphonyl derivatives were docked into the class C Enterobacter cloacae P99 Cephalosporinase. The results demonstrated that beta isomer of 4-oxo-pent-2-enyl substituent at C-6 with good bioactivity was docked deeply inside the active site and interacted well with the active site residues. However, the alpha isomer with low activity did not dock deeply inside and interacted with few active site residues. The beta isomer of 3-methoxycarbonyl-allyl substituent at C-6 which has high bioactivity was docked stably inside the active site whereas the alpha isomer with low bioactivity was partially docked. Also, the beta isomer had more hydrogen bonds with the active site residues than that of alpha isomer. (3R,5R) 3-Phenyl-clavam with the highest activity among the four diastereomers of 3-Phenyl-clavams, respectively, docked well, forming strong hydrogen bonds with the active site residues. (3S,5R) and (3S,5S) 3-Phenyl-clavams that exhibited lack of β-lactamase inhibition docked poorly with different binding modes from the (3R,5R) 3-Phenyl-clavam. (3R,5R,6S) 6-Methyl-3-Phenyl-clavam was shown best activity among four diastereomers of 6-Methyl 3-Phenyl clavams docked better into the active site than the other diastereomers. L isomer of p-nitrophenyl 1-(N-phenylacetyl amino)ethyl-phosphonate and 1-(N-benzyloxy-carbonylamino)ethyl-phosphonate with good activity was docked deeply inside the active site and interacted well with SER, key residue of mechanism, and active site residues. These results suggest that the stereoisomers can be selected by the binding mode differences through docking study. Thus, it could be a useful method to the development of therapeutically effective stereoisomers of β-lactamase inhibitors.;박테리아는 β-lactamase를 생성하여 β-lactam 항생제를 가수분해함으로써 저항성을 획득하고, 이에 대응하여 β-lactamase 저해제를 함께 사용함으로써 β-lactamase에 의한 항생제의 파괴를 막고 좋은 치료 효과를 얻을 수 있다. 그러나 β-lactamase 중 병원체들 사이에서 매우 널리 발현되고 있는 class B와 C에 속하는 효소들에 대해 기존의 저해제들은 저해작용이 약하여 이 효소들에 활성을 지닌 새로운 저해제의 개발이 시급한 실정이다. 이에 기존의 β-lactamase 저해 효과가 입증된 구조들에 여러 치환기를 도입한 새로운 화합물들이 연구, 발표 되었다. 이러한 화합물들은 입체이성체에 따라 활성이 다른 것으로 나타났는데, 이렇게 특정 입체이성체만 특이적으로 생리활성을 갖는 경우는 수용체와의 입체화학적인 상호작용에 기인하는 것으로 이해되고 있어 이성체 중 생리활성을 갖는 물질을 선별하기 위해 computer aided molecular docking이 효율적인 접근 방법으로 선택되었다. Flexible docking program인 FlexiDock과 QXP를 사용하여 sulbactam의 새로운 allyl substituted penam sulfones 유도체들의 α, β-이성체 4개는 class B의 bacteroides fragilis metallo-β-lactamase에, 3-phenyl-clavams의 4 개의 입체이성체와 6-methyl-3phenyl clavams의 4 개의 입체이성체 그리고 novel phosphonyl 유도체의 D, L 이성체 4개는 class C인 enterobacter cloacae P99 β-lactamase에 docking 하여 그 상호작용을 관찰하였다. 그 결과 allyl substituted penam sulfones 유도체 중 활성이 큰 β-이성체가 모두 active site 안 쪽으로 깊숙이 들어가 metal center에 근접하게 위치하여 active site 잔기들과 상호작용하였다. 그러나 낮은 활성을 지닌 α-이성체는 metal center에 β-이성체 만큼 접근하지 못 하였다. 3-Phenyl-clavams 중 활성이 큰 (3R5R)3-phenyl-clavams는 다른 diastereomers에 비해 더 안정적으로 docking 되었으며 active site 잔기와 강한 수소결합을 형성하였다. 반면 활성이 거의 없는 것으로 측정된 (3S,5R) 과 (3S,5S) 3- Phenyl clavams들은 active site serine에서 멀리 위치하였으며 수소결합도 형성 하지 못하였다. 한편, (3R, 5R, 6S) 6-methyl-3-phenyl-clavam들은 6-methyl-3-phenyl-clavam들 중 가장 좋은 활성을 가지고 있고 기전에서 중요한 역할을 하는 SER64에 다른 세 가지 입체이성체들 보다 근접하게 위치하며 잘 docking 되었다. Phosphonyl 유도체들 중 활성이 큰 L체는 D체에 비해 SER64과 상호작용을 잘 형성하였으며 다른 active site 잔기와의 상호작용도 많아 더 잘 결합하고 있음을 알 수 있었다. 본 실험 결과는 docking 연구에서 target enzyme과의 상호작용이 더 강하게 나타나는 입체이성체의 활성이 더 크다는 것을 보여주었다. 이러한 결과들은 docking 연구를 통한 결합 양상의 차이점을 이용해 활성이 있는 입체이성체를 선택할 수 있음을 의미한다. 따라서 computer aided molecular docking은 효과적인 β-lactamase 저해제의 개발을 위한 유용한 방법이 될 것이다.
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