The Bailinquan team of Shanghai Jiaotong University analyzed the crystal structure and catalytic mechanism of novel carbamoyltransferase

Recently, the team of Professor Bai Linquan of the School of Life Science and Technology of Shanghai Jiao Tong University published an online publication entitled "Endowing homodimeric carbamoyltransferase GdmN with iterative functions through structural characterization and" in the international authoritative journal Nature Communications Mechanistic Studies"
.
This study focused on novel dimeric carbamoyltransferases in the post-synthesis modification of ANSA drugs (amsis and geldenin), meticulously analyzed the catalytic mechanism of their proteins, and realized iterative carbamoylation remodeling
of enzyme functions.
Wei Jianhua, doctoral students of the School of Life Science and Technology, and Dr.
Zhang Xuan of Xiamen University are the first authors of the paper, and Professor Bai Linquan and Associate Researcher Kang Qianjin of the School of Life Science and Technology are the co-corresponding authors
.

Maydenin is chemically structurally a natural product of ANSA, with strong anti-tumor activity, and has been successfully used in the "warhead" molecule of antibody conjugate drugs, and drugs
for the treatment of HER2+ breast cancer have been developed.
At the same time, antibody conjugates with multiple pipelines are in the development or clinical validation stage
.
Professor Bai Linquan and several research groups have been studying the biosynthesis mechanism and metabolic pathway modification of microbial derived meltanin (ammectin) drugs for a long time, and have achieved a series of research results
.

As a class of biological macromolecules widely present in primary and secondary metabolism, carbamoyltransferases play an important role
in post-synthesis modification of ANSA drugs.
Structure-activity studies of secondary metabolites have shown that carbamoyl groups loaded by carbamoyltransferase are critical
for the biological activity of secondary metabolites.
The study found that ANSA-carbamoyltransferase GdmN forms a dimerized structure in a face-to-face form, and the special C-terminal domain realizes the function of protein dimerization through the interaction of four reverse β lamellas.
By optimizing the cocrystallization conditions of substrate and protein complex, the complex crystals of GdmN and multiple reaction intermediates were captured, the dynamic process of donor substrate molecules and acceptor substrate molecules in enzymatic reactions and key amino acid residues His27 and Tyr82 were carefully demonstrated, and the transfer mechanism
of carbamoyl groups was deeply analyzed by computational analysis.

Figure 1.
Crystal structure and catalytic mechanism of dimerized carbamoyltransferase

On this basis, the substrate recognition characteristics of dimerized carbamoyltransferase were explored by binding protein structure and phylogenetic analysis, which successfully remodeled proteins with iterative carbamoylation function, and revealed the changes of
substrate recognition position before and after protein function remodeling.

Figure 2.
Carbamoyltransferase crystal structure guided functional shaping

This study analyzed the structure of dimerized carbamoyltransferase protein, further enriched the study of carbamoyltransferase family proteins, flexibly used structural biology methods, captured the snapshots of multiple reaction intermediates in the enzyme catalytic process, combined computational biology and protein modification, successfully redesigned the active site and expanded its catalytic ability, and provided reference
for the rational transformation of other proteins.

The work was strongly supported
by Zheng Jianjian, Associate Professor Tang Mancheng, Dr.
Lin Zhi, Professor Wang Binju of Xiamen University, and Zhou Jiahai, researcher of Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences.
The work has also been supported
by the National Key Research and Development Program of China, the National Natural Science Foundation of China, and the Shanghai Municipal Science and Technology Commission.

Links to papers: