Combining oligomer build-up with alanine scanning to determine the flocculation protein mutants for enhancing oligosaccharide binding

ABSTRACT

Oligosaccharide binding on protein plays an important role in cellular signalling and recognition. Protein mutation can alter the binding strength and thus regulate the relevant processes. In this work, we investigated the interactions between the mannose oligosaccharides and the wild type and mutant forms of yeast flocculation protein Flo1p from Saccharomyces cerevisiae. The binding strength of an oligosaccharide chain was estimated using molecular dynamics simulation and MM/GBSA binding free energy calculation, via the build-up of oligosaccharide from monosaccharide to disaccharide, trisaccharide and tetrasaccharide. At the protein-disaccharide stage, several mutation positions were identified via alanine scanning mutagenesis, leading to nine mutants, from which two mutants Q117N and Q117R were selected with significantly stronger binding free energies relative to the wild type protein. Furthermore, the two mutants also show enhanced binding at the trisaccharide and tetrasaccharide stages. Conformational stability and shape change were analysed to help understand the protein-oligosaccharide interactions. This study also presented a computational example to estimate the oligosaccharide binding strength using its terminal sugar residues.

KEYWORDS:

• Alanine scanning mutagenesis
• binding free energy
• molecular dynamics simulations
• oligosaccharide binding
• yeast flocculation protein mutant

Disclosure statement

No potential conflict of interest was reported by the author(s).

Authors’ contributions

Sun did the most computations. Zhang did the primary computations which lead to the conclusions. Chen did the auxiliary computations. All of the above three authors prepared the figures and tables. Zhao, Bai and Zhong designed the research plan. Sun, Zhang and Zhong wrote the manuscript.

Additional information

Funding

This work was supported by the National Natural Science Foundation of China (No. 21536006). We thank Christoffer Lind and Weixi Zhong for helpful discussions.