the Vmax/Km value of the enzyme for cis-4-hydroxy-D-proline is about 124fold and about 736fold higher than the Vmax/Km values for trans-4-hydroxy-D-proline and D-proline, respectively. Both AbLhpBEF (heteromer of alpha-, beta- and gamma-subunit) and AbLhpB (catalytic beta-subunit) are strictly specific to cis-4-hydroxy-D-proline
the Vmax/Km value of the enzyme for cis-4-hydroxy-D-proline is about 124fold and about 736fold higher than the Vmax/Km values for trans-4-hydroxy-D-proline and D-proline, respectively. Both AbLhpBEF (heteromer of alpha-, beta- and gamma-subunit) and AbLhpB (catalytic beta-subunit) are strictly specific to cis-4-hydroxy-D-proline
the Vmax/Km value of the enzyme for cis-4-hydroxy-D-proline is about 124fold and about 736fold higher than the Vmax/Km values for trans-4-hydroxy-D-proline and D-proline, respectively. Both AbLhpBEF (heteromer of alpha-, beta- and gamma-subunit) and AbLhpB (catalytic beta-subunit) are strictly specific to cis-4-hydroxy-D-proline
alpha4beta4gamma4, the gamma-subunit significantly contributes to stabilizing the alpha4beta4gamma4 structure (and the alpha-subunit), but not to enzyme catalysis
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CLONED/commentary
ORGANISM
UNIPROT
LITERATURE
cloning of the AbLhpB, AbLhpE and AbLhpF genes into different plasmid vectors, construction of a functional expression system of recombinant HypDH with a heterooligomeric structure in Escherichia coli cells
estimation of the content of L-hydroxyprolines using coupling systems with metabolic enzymes of the trans-4-hydroxy-L-proline pathway (hydroxyproline 2-epimerase (HypE) and cis-4-hydroxy-D-proline dehydrogenase (HypDH)) and the trans-3-hydroxy-L-proline pathway (trans-3-hydroxy-L-proline dehydratase (T3LHypD) and DELTA1-pyrroline-2-carboxylate reductase (Pyr2CR)) from microorganisms. A functional expression system of recombinant HypDH with a heterooligomeric structure is constructed in Escherichia coli cells. Enzymological characterization reveals that the beta-subunit acts as a catalytic subunit, and also that assembly with other subunit(s) improves the kinetics for cis-4-hydroxy-D-proline and thermostability. By using a spectrophotometric assay with different wavelengths, the contents of trans-4-hydroxy-L-proline and trans-3-hydroxy-L-proline are successfully estimated within the ranges of 0.004-1 mM and 0.05-1 mM, respectively, and are consistent with the contents determined by HPLC. This enzymatic method is used to measure the content of trans-4-hydroxy-L-proline in the acid-hydrolysate of collagen, and blood plasma
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1.4.99.B4 cis-4-hydroxy-D-proline dehydrogenase
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