PDF(Pubmed)
Abstract：
Lactate dehydrogenase A (LDHA) primarily catalyzes the conversion between lactic acid and pyruvate, serving as a key enzyme in the aerobic glycolysis pathway of sugar in tumor cells. LDHA plays a crucial role in the occurrence, development, progression, invasion, metastasis, angiogenesis, and immune escape of tumors. Consequently, LDHA not only serves as a biomarker for tumor diagnosis and prognosis but also represents an ideal target for tumor therapy. Although LDHA inhibitors show great therapeutic potential, their development has proven to be challenging. In the development of LDHA inhibitors, the key active sites of LDHA are emphasized. Nevertheless, there is a relative lack of research on the amino acid residues around the active center of LDHA. Therefore, in this study, we investigated the amino acid residues around the active center of LDHA. Through structure comparison analysis, five key amino acid residues (Ala30, Met41, Lys131, Gln233, and Ala259) were identified. Subsequently, the effects of these five residues on the enzymatic properties of LDHA were investigated using site-directed mutagenesis. The results revealed that the catalytic activities of the five mutants varied to different degrees in both the reaction from lactic acid to pyruvate and pyruvate to lactic acid. Notably, the catalytic activities of LDHAM41G and LDHAK131I were improved, particularly in the case of LDHAK131I. The results of the molecular dynamics analysis of LDHAK131I explained the reasons for this phenomenon. Additionally, the optimum temperature of LDHAM41G and LDHAQ233M increased from 35 °C to 40 °C, whereas in the reverse reaction, the optimum temperature of LDHAM41G and LDHAK131I decreased from 70 °C to 60 °C. These findings indicate that Ala30, Met41, Lys131, Gln233, and Ala259 exert diverse effects on the catalytic activity and optimum temperature of LHDA. Therefore, these amino acid residues, in addition to the key catalytic site of the active center, play a crucial role. Considering these residues in the design and screening of LDHA inhibitors may lead to the development of more effective inhibitors.
摘要：
乳酸脱氢酶A（LDHA）主要催化乳酸和丙酮酸之间的转化，作为肿瘤细胞中糖的有氧糖酵解途径的关键酶。LDHA在其发生中起着至关重要的作用,发展,programming,入侵,转移,血管生成,和肿瘤的免疫逃逸。因此,LDHA不仅是肿瘤诊断和预后的生物标志物，也是肿瘤治疗的理想靶点。虽然LDHA抑制剂显示出巨大的治疗潜力，事实证明，他们的发展具有挑战性。在LDHA抑制剂的开发中，强调了LDHA的关键活性位点。然而，对LDHA活性中心周围氨基酸残基的研究相对缺乏。因此,在这项研究中，我们研究了LDHA活性中心周围的氨基酸残基。通过结构对比分析,鉴定了五个关键氨基酸残基(Ala30、Met41、Lys131、Gln233和Ala259)。随后，使用定点诱变研究了这五个残基对LDHA酶学性质的影响。结果表明，从乳酸到丙酮酸和丙酮酸到乳酸的反应中，五种突变体的催化活性都有不同程度的变化。值得注意的是,LDHAM41G和LDHAK131I的催化活性得到了提高,特别是在LDHAK131I的情况下。LDHAK131I的分子动力学分析结果解释了这种现象的原因。此外,LDHAM41G和LDHAQ233M的最佳温度从35°C增加到40°C，而在逆反应中，LDHAM41G和LDHAK131I的最佳温度从70℃降至60℃。这些发现表明Ala30，Met41，Lys131，Gln233和Ala259对LHDA的催化活性和最佳温度具有不同的影响。因此,这些氨基酸残基，除了活性中心的关键催化位点，发挥关键作用。在LDHA抑制剂的设计和筛选中考虑这些残基可能导致开发更有效的抑制剂。
