# NMR study of the structure and dynamics of the BRCT domain from the kinetochore protein KKT4

## TL;DR

This study uses NMR to analyze the BRCT domain of the KKT4 protein in Trypanosoma brucei, revealing its structure and phosphate-binding function important for parasite growth.

## Contribution

The study provides NMR resonance assignments and functional insights into the BRCT domain of KKT4, a protein unique to kinetoplastids.

## Key findings

- The BRCT domain of KKT4 binds phosphate ions in solution using residues that also bind sulfate ions in the X-ray structure.
- Mutations in phosphate-binding residues of the BRCT domain cause growth defects in Trypanosoma brucei.
- NMR resonance assignments help characterize the secondary structure and dynamics of the BRCT domain in solution.

## Abstract

KKT4 is a multi-domain kinetochore protein specific to kinetoplastids, such as Trypanosoma brucei. It lacks significant sequence similarity to known kinetochore proteins in other eukaryotes. Our recent X-ray structure of the C-terminal region of KKT4 shows that it has a tandem BRCT (BRCA1 C Terminus) domain fold with a sulfate ion bound in a typical binding site for a phosphorylated serine or threonine. Here we present the 1H, 13C and 15N resonance assignments for the BRCT domain of KKT4 (KKT4463–645) from T. brucei. We show that the BRCT domain can bind phosphate ions in solution using residues involved in sulfate ion binding in the X-ray structure. We have used these assignments to characterise the secondary structure and backbone dynamics of the BRCT domain in solution. Mutating the residues involved in phosphate ion binding in T. brucei KKT4 BRCT results in growth defects confirming the importance of the BRCT phosphopeptide-binding activity in vivo. These results may facilitate rational drug design efforts in the future to combat diseases caused by kinetoplastid parasites.

The online version contains supplementary material available at 10.1007/s12104-024-10163-9.