Glutamate 1-semialdehyde aminotransferase (GSA-AT) is a key regulatory enzyme, which converts glutamate 1-semialdehyde (GSA) to 5-aminolevulinic acid (ALA) in chlorophyll biosynthesis. ALA is the universal precursor for the synthesis of chlorophyll, heme, and other tetrapyrroles. To study the regulation of chlorophyll biosynthesis in Brassica napus, two cDNA clones of GSA-AT were isolated for genetic manipulation. A SalI-XbaI fragment from one of the two cDNA clones of GSA-AT was used for recombinant protein expression by inserting it at the 3' end of a calmodulin-binding-peptide (CBP) tag of the pCaln vector. The CBP tagged recombinant protein, expressed in Escherichia coli, was purified to apparent homogeneity in a one step purification process using a calmodulin affinity column. The purified CBP tagged GSA-AT is biologically active and has a specific activity of 16.6 nmol/min/mg. Cleavage of the CBP tag from the recombinant protein with thrombin resulted in 9.2% loss of specific activity. However, removal of the cleaved CBP tag from the recombinant protein solution resulted in 60% loss of specific activity, suggesting possible interactions between the recombinant protein and the CBP tag. The enzyme activity of the CBP tagles...Continue Reading
A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding
The role of Lys272 in the pyridoxal 5-phosphate active site of Synechococcus glutamate-1-semialdehyde aminotransferase
Activity and spectroscopic properties of the Escherichia coli glutamate 1-semialdehyde aminotransferase and the putative active site mutant K265R
Characterization of glutamate-1-semialdehyde aminotransferase of Synechococcus. Steady-state kinetic analysis
Conditions for the assay of glutamate semialdehyde aminotransferase that overcome the problem of substrate instability
Structural genes of glutamate 1-semialdehyde aminotransferase for porphyrin synthesis in a cyanobacterium and Escherichia coli
Cloning and sequence of the Salmonella typhimurium hemL gene and identification of the missing enzyme in hemL mutants as glutamate-1-semialdehyde aminotransferase
Primary structure of a key enzyme in plant tetrapyrrole synthesis: glutamate 1-semialdehyde aminotransferase
Purification and partial amino acid sequence of the glutamate 1-semialdehyde aminotransferase of barley and synechococcus
tRNA(Glu) as a cofactor in delta-aminolevulinate biosynthesis: steps that regulate chlorophyll synthesis
Reactions of glutamate semialdehyde aminotransferase (glutamate-1-semialdehyde 2,1 aminomutase) with vinyl and acetylenic substrate analogues analysed by rapid scanning spectrophotometry
Light regulation of chlorophyll biosynthesis at the level of 5-aminolevulinate formation in Arabidopsis
Crystallization and preliminary X-ray analysis of wild-type and K272A mutant glutamate 1-semialdehyde aminotransferase from Synechococcus
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Expression of the soybean (Glycine max) glutamate 1-semialdehyde aminotransferase gene in symbiotic root nodules
Properties of the pyridoxaldimine form of glutamate semialdehyde aminotransferase (glutamate-1-semialdehyde 2,1-aminomutase) and analysis of its role as an intermediate in the formation of aminolaevulinate
Chlorophyll reduction in the seed of Brassica napus with a glutamate 1-semialdehyde aminotransferase antisense gene
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Purification and Characterization of Glutamate 1-Semialdehyde Aminotransferase from Barley Expressed in Escherichia coli.
Identification and characterization of a pyridoxal reductase involved in the vitamin B6 salvage pathway in Arabidopsis.
Cellular levels of glutamyl-tRNA reductase and glutamate-1-semialdehyde aminotransferase do not control chlorophyll synthesis in Chlamydomonas reinhardtii
Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of the glutamate-1-semialdehyde aminotransferase from Bacillus subtilis.
A Single Nucleotide Substitution of GSAM Gene Causes Massive Accumulation of Glutamate 1-Semialdehyde and Yellow Leaf Phenotype in Rice.
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