Mining for allosteric information: natural mutations and positional sequence conservation in pyruvate kinase

Area of Science:

• Biochemistry
• Molecular Biology
• Enzymology

Background:

• Pyruvate kinase (PYK) isozymes share conserved structures and sequences.
• Allosteric regulation mechanisms vary significantly among PYK isozymes.
• This suggests a role for less conserved amino acids in allosteric control.

Purpose of the Study:

• To identify amino acid positions critical for pyruvate kinase (PYK) allosteric regulation.
• To leverage naturally occurring mutations associated with nonspherocytic hemolytic anemia as a 'natural screen'.
• To correlate mutation data with amino acid conservation across PYK sequences.

Main Methods:

• Analysis of 122 point mutations in the human erythrocyte PYK isozyme gene linked to hemolytic anemia.
• Comparison of mutation data with amino acid conservation across 241 PYK sequences.
• Categorization of residue positions into three groups based on mutation presence and conservation levels.

Main Results:

• Mutations at highly conserved positions likely impair fundamental PYK functions (structure, catalysis).
• Mutations at non-conserved positions, identified in disease states, are candidates for allosteric roles.
• Group 3 residues (mutated and low conservation) are implicated in isozyme-specific functions like allostery, often located in C and A domains.

Conclusions:

• Amino acid positions with low conservation and identified disease-causing mutations are crucial for PYK allosteric regulation.
• This study identifies specific residues likely responsible for differential allosteric control among PYK isozymes.
• Findings highlight the importance of non-conserved residues in enzyme function beyond basic catalysis and structure.