Interaction between pyridoxal kinase and pyridoxal-5-phosphate-dependent enzymes.
The interactions of two pyridoxal-5-phosphate (PLP)-dependent enzymes, alanine aminotransferase (ALT) and glutamate decarboxylase (GAD), with pyridoxal kinase (PK) were studied by fluorescence polarization as well as surface plasmon resonance techniques. The results demonstrated that PK can specifically bind to ALT and GAD. Moreover, binding profiles of both enzymes to immobilized PK were altered by excess amount of PLP. The equilibrium affinity constants for ALT in the absence and presence of PLP are 20.4 x 10(4) M(-1)and 6.7 x 10(4) M(-1), and for GAD are 37 x 10(4) M(-1)and 20.8 x 10(4) M(-1), respectively. It appears that specific interactions occur between PK and PLP-dependent enzymes, and the binding affinities of PK for PLP-dependent enzymes decrease in the presence of PLP. The results support our hypothesis that PLP transfer from PK to PLP-dependent enzymes requires a specific interaction between PK and the enzyme.
Cheung
Pik-Yuen
PY
The Central Laboratory of the Institute of Molecular Technology for Drug Discovery and Synthesis, Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University, Hung Hom, Kowloon, Hong Kong SAR, P.R.C.
Fong
Chi-Chun
CC
Ng
Kang-To
KT
Lam
Wan-Chuen
WC
Leung
Yun-Chung
YC
Tsang
Chun-Wai
CW
Yang
Mengsu
M
Wong
Man-Sau
MS
D000410:Alanine Transaminase / Q000378:metabolism*; D000818:Animals; D004800:Enzymes, Immobilized / Q000378:metabolism; D005454:Fluorescence Polarization; D005968:Glutamate Decarboxylase / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D011485:Protein Binding; D011731:Pyridoxal Kinase / Q000302:isolation & purification / Q000378:metabolism*; D011732:Pyridoxal Phosphate / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013552:Swine; D013997:Time Factors
D004800:Enzymes, Immobilized; D011994:Recombinant Proteins; D011732:Pyridoxal Phosphate; D000410:Alanine Transaminase; D011731:Pyridoxal Kinase; D005968:Glutamate Decarboxylase