PMID:12611890

Citation	Pezza, JA, Choi, KH, Berardini, TZ, Beernink, PT, Allen, KN and Tolan, DR (2003) Spatial clustering of isozyme-specific residues reveals unlikely determinants of isozyme specificity in fructose-1,6-bisphosphate aldolase. J. Biol. Chem. 278:17307-13
Abstract	Vertebrate fructose-1,6-bisphosphate aldolase exists as three isozymes (A, B, and C) that demonstrate kinetic properties that are consistent with their physiological role and tissue-specific expression. The isozymes demonstrate specific substrate cleavage efficiencies along with differences in the ability to interact with other proteins; however, it is unknown how these differences are conferred. An alignment of 21 known vertebrate aldolase sequences was used to identify all of the amino acids that are specific to each isozyme, or isozyme-specific residues (ISRs). The location of ISRs on the tertiary and quaternary structures of aldolase reveals that ISRs are found largely on the surface (24 out of 27) and are all outside of hydrogen bonding distance to any active site residue. Moreover, ISRs cluster into two patches on the surface of aldolase with one of these patches, the terminal surface patch, overlapping with the actin-binding site of aldolase A and overlapping an area of higher than average temperature factors derived from the x-ray crystal structures of the isozymes. The other patch, the distal surface patch, comprises an area with a different electrostatic surface potential when comparing isozymes. Despite their location distal to the active site, swapping ISRs between aldolase A and B by multiple site mutagenesis on recombinant expression plasmids is sufficient to convert the kinetic properties of aldolase A to those of aldolase B. This implies that ISRs influence catalysis via changes that alter the structure of the active site from a distance or via changes that alter the interaction of the mobile C-terminal portion with the active site. The methods used in the identification and analysis of ISRs discussed here can be applied to other protein families to reveal functionally relevant residue clusters not accessible by conventional primary sequence alignment methods.
Links	PubMed Online version:10.1074/jbc.M209185200
Keywords	Amino Acid Sequence; Animals; Chickens; Enzyme Stability; Fructose-Bisphosphate Aldolase/chemistry; Fructose-Bisphosphate Aldolase/genetics; Fructose-Bisphosphate Aldolase/metabolism; Humans; Isoenzymes/chemistry; Isoenzymes/genetics; Isoenzymes/metabolism; Kinetics; Mice; Molecular Sequence Data; Organ Specificity; Protein Conformation; Rabbits; Rats; Recombinant Proteins/chemistry; Recombinant Proteins/genetics; Recombinant Proteins/metabolism; Sequence Alignment; Substrate Specificity; Xenopus
edit table

Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
MYCTU:ALF		GO:0042132: fructose 1,6-bisphosphate 1-phosphatase activity	ECO:0000315:		F		The kinetic constants for recombinant wild-type aldolase B were obtained in the same fashion as for the mutant enzymes shown in Table 2.	complete CACAO 5206

References

See Help:References for how to manage references in GONUTS.