School of Chemistry and the Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, 30 Flemington Rd, Parkville, VIC 3052, Australia.
School of Chemical Sciences, The University of Auckland, Private Bag 92019, Auckland, 1142, New Zealand.
Chembiochem. 2023 Jul 17;24(14):e202300162. doi: 10.1002/cbic.202300162. Epub 2023 Jun 18.
Isocitrate lyase (ICL) isoform 2 is an essential enzyme for some clinical Mycobacterium tuberculosis (Mtb) strains during infection. In the laboratory Mtb strain H37Rv, the icl2 gene encodes two distinct gene products - Rv1915 and Rv1916 - due to a frameshift mutation. This study aims to characterise these two gene products to understand their structure and function. While we were unable to produce Rv1915 recombinantly, soluble Rv1916 was obtained with sufficient yield for characterisation. Kinetic studies using UV-visible spectrophotometry and H-NMR spectroscopy showed that recombinant Rv1916 does not possess isocitrate lyase activity, while waterLOGSY binding experiments demonstrated that it could bind acetyl-CoA. Finally, X-ray crystallography revealed structural similarities between Rv1916 and the C-terminal domain of ICL2. Considering the probable differences between full-length ICL2 and the gene products Rv1915 and Rv1916, care must be taken when using Mtb H37Rv as a model organism to study central carbon metabolism.
异柠檬酸裂解酶 (ICL) 同工酶 2 是一些临床结核分枝杆菌 (Mtb) 菌株在感染过程中的必需酶。在实验室 Mtb 菌株 H37Rv 中，icl2 基因由于移码突变而编码两个不同的基因产物 - Rv1915 和 Rv1916。本研究旨在对这两个基因产物进行表征，以了解它们的结构和功能。虽然我们无法重组产生 Rv1915，但获得了足够用于表征的可溶性 Rv1916。使用紫外-可见分光光度法和 H-NMR 光谱法进行的动力学研究表明，重组 Rv1916 不具有异柠檬酸裂解酶活性，而水 LOGSY 结合实验表明它可以结合乙酰辅酶 A。最后，X 射线晶体学揭示了 Rv1916 和 ICL2 的 C 末端结构域之间的结构相似性。考虑到全长 ICL2 与基因产物 Rv1915 和 Rv1916 之间可能存在的差异，在使用 Mtb H37Rv 作为研究中心碳代谢的模式生物时，必须小心谨慎。
