Verification of the Stabilized Protein Design Based on the Prediction of Intrinsically Disordered Regions: Ribosomal Proteins L1

Biochemistry (Mosc). 2020 Jan;85(1):90-98. doi: 10.1134/S0006297920010083.

Abstract

In our previous papers, we proposed the idea that programs predicting intrinsically disordered regions in amino acid sequences can be used for finding weakened sites in proteins. The regions predicted by such programs are suitable targets for the introduction of protein-stabilizing mutations. However, for each specific protein, it remains unclear what determines protein stabilization - the amino acid sequence (and accordingly, prediction of weakened sites) or the 3D structure. To answer this question, it is necessary to study two proteins with similar structures but different amino acid sequences and, consequently, different predictions of weakened regions. By introducing identical mutations into identical elements of the two proteins, we will be able to reveal whether predictions of the weakened sites or the 3D protein structure are the key factors in the protein stability increase. Here, we have chosen ribosomal proteins L1 from the halophilic archaeon Haloarcula marismortui (HmaL1) and extremophilic bacterium Aquifex aeolicus (AaeL1). These proteins are identical in their structure but different in amino acid sequences. A disulfide bond introduced into the region predicted as the structured one in AaeL1 did not lead to the increase in the protein melting temperature. At the same time, a disulfide bond introduced into the same region in HmaL1 that was predicted as a weakened one, resulted in the increase in the protein melting temperature by approximately 10°C.

MeSH terms

  • Amino Acid Sequence
  • Aquifex
  • Archaeal Proteins / chemistry*
  • Bacteria / metabolism*
  • Bacterial Proteins / chemistry*
  • Cloning, Molecular
  • Escherichia coli / genetics
  • Haloarcula marismortui / metabolism*
  • Models, Molecular
  • Protein Stability
  • Protein Structure, Tertiary
  • Ribosomal Proteins / chemistry*

Substances

  • Archaeal Proteins
  • Bacterial Proteins
  • Ribosomal Proteins
  • ribosomal protein L1

Supplementary concepts

  • Aquifex aeolicus