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

Abstract

Although the amino acid sequences and the structures of pyruvate kinase (PYK) isozymes are highly conserved, allosteric regulations differ. This suggests that amino acids with low conservation play important roles in the allosteric mechanism. The current work exploits a 'natural screen'‐ the 122 point mutations identified in the human gene encoding the erythrocyte PYK isozyme and associated with nonspherocytic hemolytic anemia ‐ to learn what amino acid positions in PYK may be important for allosteric regulations. In addition to the mutations, we consider the conservation of each amino acid position across 241 PYK sequences. Three groups of residue positions have been created, those with: (1) no disease causing mutation identified; (2) a disease causing mutation identified and high conservation across isozymes; and (3) a disease causing mutation identified and low conservation. Mutations at positions not identified in the natural screen are likely to be tolerated with minimal loss of function. Mutations at highly conserved positions are more likely to disrupt properties common to all PYK isozymes (e.g., structure, catalysis). Residues in the third group are likely to be involved in roles that are necessary for function but not common to all isozymes (e.g., allostery). Many of the Group 3 residues are located in the C‐domain and to a lesser extent the A domain. IUBMB Life, 58: 31 ‐ 38, 2006

Sulphate Removal Induces a Major Conformational Change in Leishmania mexicana Pyruvate Kinase in the Crystalline State

“…The site point mutation data presented here for LmPYK and for a number of other PYKs 25 show that profound changes in enzymatic activity can be achieved by a broad variety of mutants remote from the active site or the effector site. In the above example, mutations in the large interface are some 30 Å away from the binding site where the effects are observed.…”

“…Of potential medical interest are the 122 naturally occurring mutations that have been identified in human erythrocyte PYK and are linked to the disease nonspherocytic haemolytic anemia. 25 A number of these mutations, occurring mainly in the C domain, are not highly conserved over the 241 species and isoforms examined and are therefore hypothesised to play a role in allostery.…”

Allosteric Mechanism of Pyruvate Kinase from Leishmania mexicana Uses a Rock and Lock Model

“…The 6 o rigid body rocking motion (Fig. 2e) of LmPYK moves a crucially important Arg 310 side chain (mutation of the equivalent Arg in E. coli PYK results in total inactivity (32), and a mutation to either Lys or Trp results in PYK deficiency in humans (19)) into the vicinity of the active site of the adjacent subunit (Fig. 2b), where it forms two stabilizing hydrogen bonds with the backbone carbonyls (residues Arg 262 and Gly 263 ) belonging to the oxalate (phosphoenolpyruvate)-stabilized A␣6Ј helix.…”

A Critical Evaluation of Correlated Mutation Algorithms and Coevolution Within Allosteric Mechanisms

“…For example, as discussed by del Sol et al (13), the plasticity within an allosteric response is such that nearby residues can easily functionally substitute for one another. Related, it has been shown in several systems (e.g., pyruvate kinase (59)) that mutations at highly variable positions distal to the active site can have dramatic impacts on catalytic activity. As such, based on the ubiquitous diversity of allosteric mechanisms, Kuriyan and Eisenberg (43) have intriguingly suggested that allosteric diversity is responsible for the complexity of life.…”