PMID- 3038027
OWN - NLM
STAT- MEDLINE
DCOM- 19870827
LR  - 20190628
IS  - 0003-9861 (Print)
IS  - 0003-9861 (Linking)
VI  - 256
IP  - 1
DP  - 1987 Jul
TI  - Spinach leaf ribulose-5-phosphate kinase: examination of sulfhydryls by chemical 
      modification and spin-labeling.
PG  - 362-71
AB  - Methodology has been developed for complete or selective modification of the 
      cysteinyl sulfhydryls of ribulose-5-phosphate (Ru5P) kinase. Using native enzyme, 
      iodoacetate modifies four sulfhydryls with varying levels of completeness. The 
      most reactive sulfhydryl in the native enzyme can be selectively titrated with 
      iodoacetate; complete loss of activity occurs. Composition and N-terminal 
      analyses of the peptide bearing this essential sulfhydryl indicate that the 
      alkylated residue (Cys-16) is identical to the site modified by other 
      modification reagents (M. A. Porter and F. C. Hartman (1986) Biochemistry 25, 
      7314-7318). In the presence of ATP, a nonessential sulfhydryl of the native 
      enzyme is carboxymethylated. The peptide bearing this modified cysteine has been 
      isolated and its composition and N-terminal sequence determined. Enzyme that is 
      carboxymethylated in the presence of ATP retains activity and can be oxidatively 
      inactivated in a reversible fashion. This suggests that the cysteine targeted by 
      iodoacetate in the presence of ATP is not a residue that participates in 
      regulation of enzyme activity. Using a spin-labeled analog of iodoacetate, both 
      essential and nonessential cysteines have been selectively modified. ESR 
      measurements suggest that the environment of these cysteines is not highly 
      constrained. Modest effects on spin-label mobility are observed upon occupancy of 
      Ru5P or ATP sites on the modified enzyme. These effects are dependent on the 
      presence of divalent cations, suggesting that a binary enzyme-cation complex must 
      form prior to productive enzyme-substrate interactions.
FAU - Krieger, T J
AU  - Krieger TJ
FAU - Miziorko, H M
AU  - Miziorko HM
LA  - eng
PT  - Journal Article
PT  - Research Support, U.S. Gov't, Non-P.H.S.
PL  - United States
TA  - Arch Biochem Biophys
JT  - Archives of biochemistry and biophysics
JID - 0372430
RN  - 0 (Amino Acids)
RN  - 0 (Iodoacetates)
RN  - 0 (Spin Labels)
RN  - 0 (Sulfhydryl Compounds)
RN  - EC 2.7.- (Phosphotransferases)
RN  - EC 2.7.1.- (Phosphotransferases (Alcohol Group Acceptor))
RN  - EC 2.7.1.19 (phosphoribulokinase)
RN  - K848JZ4886 (Cysteine)
RN  - WF5188V710 (Iodoacetic Acid)
SB  - IM
MH  - Amino Acids/analysis
MH  - Chromatography/methods
MH  - Cysteine/analysis
MH  - Electron Spin Resonance Spectroscopy
MH  - Iodoacetates
MH  - Iodoacetic Acid
MH  - Peptide Mapping
MH  - *Phosphotransferases
MH  - *Phosphotransferases (Alcohol Group Acceptor)
MH  - Plants/*enzymology
MH  - Spin Labels
MH  - Sulfhydryl Compounds/*analysis
EDAT- 1987/07/01 00:00
MHDA- 1987/07/01 00:01
CRDT- 1987/07/01 00:00
PHST- 1987/07/01 00:00 [pubmed]
PHST- 1987/07/01 00:01 [medline]
PHST- 1987/07/01 00:00 [entrez]
AID - 0003-9861(87)90457-7 [pii]
AID - 10.1016/0003-9861(87)90457-7 [doi]
PST - ppublish
SO  - Arch Biochem Biophys. 1987 Jul;256(1):362-71. doi: 10.1016/0003-9861(87)90457-7.