Jena Lingaraja, Deshmukh Shraddha, Waghmare Pranita, Kumar Satish, Harinath Bhaskar C
Bioinformatics Centre, Biochemistry and JB Tropical Disease Research Centre, Mahatma Gandhi Institute of Medical Sciences, Sevagram, Maharashtra, India.
Bioinformatics Centre, Biochemistry and JB Tropical Disease Research Centre, Mahatma Gandhi Institute of Medical Sciences, Sevagram, Maharashtra, India.
Int J Mycobacteriol. 2015 Dec;4(4):276-83. doi: 10.1016/j.ijmyco.2015.06.006. Epub 2015 Jul 15.
OBJECTIVE/BACKGROUND: Isoniazid (INH) is one of the effective antituberculosis (TB) drugs used for TB treatment. However, most of the drug-resistant Mycobacterium tuberculosis (MTB) clinical strains are resistant to INH, a first-line antituberculous drug. Certain metabolic enzymes such as adenosylhomocysteinase (Rv3248c), universal stress protein (Rv2623), nicotinamide adenine dinucleotide (reduced)-dependent enoyl-acyl carrier protein reductase (Rv1484), oxidoreductase (Rv2971), dihydrofolate reductase (Rv2763c), pyrroline-5-carboxylate dehydrogenase (Rv1187) have been identified to bind INH-nicotinamide adenine dinucleotide (INH-NAD) and INH-nicotinamide adenine dinucleotide phosphate adducts coupled to Sepharose resin. These enzymes are reported to be involved in many important biochemical processes of MTB, including cysteine and methionine metabolism, mycobacterial growth regulation, mycolic acid biosynthesis, detoxification of toxic metabolites, folate biosynthesis, etc. The truncated INH-nicotinamide adenine dinucleotide (oxidized) adduct, 4-isonicotinoylnicotinamide, isolated from urine samples of human TB patients treated with INH therapy is proposed to have antimycobacterial activity.
To understand the mechanism of interaction of the truncated INH-NAD adduct, binding energy studies were carried out on the aforementioned six enzymes with known three-dimensional structures using AutoDock4.2.
In silico docking analysis of these MTB enzymes with the truncated INH-NAD adduct showed favorable binding interactions with docking energies ranging from -5.29 to -7.07 kcal/mol.
Thus, in silico docking study revealed that the INH-NAD adduct, which is generated in vivo after INH activation, may undergo spontaneous hydrolysis to form the truncated INH-NAD adduct and further binds and inhibits multiple enzymes of MTB, in addition to InhA, confirming that INH is an effective anti-TB drug acting at multiple enzymes. Further analysis of amino acid residues in the active site of INH-NAD-binding proteins showed the probable presence of catalytic triad in four enzymes possibly involved in INH binding to the enzyme.
目的/背景：异烟肼（INH）是用于结核病治疗的有效抗结核药物之一。然而，大多数耐药结核分枝杆菌（MTB）临床菌株对一线抗结核药物异烟肼耐药。已确定某些代谢酶，如腺苷同型半胱氨酸酶（Rv3248c）、通用应激蛋白（Rv2623）、烟酰胺腺嘌呤二核苷酸（还原型）依赖性烯酰 - 酰基载体蛋白还原酶（Rv1484）、氧化还原酶（Rv2971）、二氢叶酸还原酶（Rv2763c）、吡咯啉 - 5 - 羧酸脱氢酶（Rv1187），可与结合到琼脂糖树脂上的异烟肼 - 烟酰胺腺嘌呤二核苷酸（INH - NAD）和异烟肼 - 烟酰胺腺嘌呤二核苷酸磷酸加合物结合。据报道，这些酶参与MTB的许多重要生化过程，包括半胱氨酸和蛋氨酸代谢、分枝杆菌生长调节、分枝菌酸生物合成、有毒代谢产物解毒、叶酸生物合成等。从接受异烟肼治疗的人类结核病患者尿液样本中分离出的截短型异烟肼 - 烟酰胺腺嘌呤二核苷酸（氧化型）加合物4 - 异烟酰烟酰胺被认为具有抗分枝杆菌活性。
为了解截短型INH - NAD加合物的相互作用机制，使用AutoDock4.2对上述六种具有已知三维结构的酶进行了结合能研究。
这些MTB酶与截短型INH - NAD加合物的计算机模拟对接分析显示出良好的结合相互作用，对接能量范围为 - 5.29至 - 7.07 kcal/mol。
因此，计算机模拟对接研究表明，异烟肼活化后在体内产生的INH - NAD加合物可能会自发水解形成截短型INH - NAD加合物，并进一步结合并抑制MTB的多种酶（除InhA外），证实异烟肼是一种作用于多种酶的有效抗结核药物。对INH - NAD结合蛋白活性位点氨基酸残基的进一步分析表明，四种可能参与异烟肼与酶结合的酶中可能存在催化三联体。
