Rat aldolase isozyme gene.

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Aldolase B is a key enzyme in glycolysis and gluconeogenesis.
• Understanding the genetic sequence of aldolase B is crucial for studying its function and related metabolic disorders.

Purpose of the Study:

• To construct and characterize complementary DNA (cDNA) clones representing the rat aldolase B gene.
• To elucidate the nucleotide sequence and identify key functional regions of the aldolase B mRNA.

Main Methods:

• Partial purification of rat aldolase B messenger RNA (mRNA) from liver polysomes.
• Synthesis of double-stranded cDNA and insertion into the pBR322 plasmid vector.
• Colony hybridization and hybrid selection-translation assays for clone screening.
• Nucleotide sequencing and restriction mapping of cDNA inserts.

Main Results:

• Isolation of recombinant plasmids with aldolase B cDNA inserts ranging from 600 to 1000 base pairs.
• Construction of a contiguous cDNA sequence spanning 1200 base pairs, representing approximately 70% of the aldolase B mRNA.
• Determination of the 3' untranslatable region and the sequence of 180 amino acids from the COOH terminus.
• Identification of a putative active site peptide with homology to other aldolase enzymes.

Conclusions:

• The generated cDNA clones provide a significant portion of the rat aldolase B gene sequence.
• The sequence data offers insights into the protein's C-terminal structure and active site.
• This work facilitates further investigations into aldolase B gene expression and function.