PMID- 3018082
OWN - NLM
STAT- MEDLINE
DCOM- 19861015
LR  - 20161123
IS  - 0022-1767 (Print)
IS  - 0022-1767 (Linking)
VI  - 137
IP  - 6
DP  - 1986 Sep 15
TI  - Characterization of the plasma membrane bound GTPase from rabbit neutrophils. I. 
      Evidence for an Ni-like protein coupled to the formyl peptide, C5a, and 
      leukotriene B4 chemotaxis receptors.
PG  - 1961-70
AB  - We have characterized the GTPase activity of the Ni-like 
      guanine-nucleotide-binding regulatory protein in rabbit neutrophil plasma 
      membranes. The low Km (3.64 +/- 0.87 X 10(-7) M) GTPase copurified with the 
      formyl peptide receptor in the plasma membrane fraction obtained by discontinuous 
      sucrose density gradient centrifugation. The Vmax (23.9 +/- 2.91 pmol/mg/min) and 
      Km of the unstimulated enzyme were similar to those reported for Ni in other cell 
      types. The activity of the unstimulated enzyme was both magnesium and sodium 
      dependent and linear over the first 4 min of the assay. The chemoattractants, 
      formyl-methionyl-leucyl-phenylalanine (fMLP), C5a, and leukotriene B4 (LTB4) 
      stimulated the GTPase in purified neutrophil plasma membrane preparations, 
      whereas other secretagogues, such as A23187 and PMA, were without effect. 
      Lineweaver-Burk analysis showed an fMLP-induced increase in Vmax (31.94 +/- 4.80 
      pmol/mg/min) (33.1 +/- 9.5%) but not in Km. The dose-response curve for fMLP 
      stimulation showed an ED50 of 4.1 +/- 1.0 X 10(-8) M and an overall 22.2 +/- 3.1% 
      maximal stimulation. C5a (30 micrograms/ml) increased the activity of the GTPase 
      21.3 +/- 5.7% and 10(-7) M LTB4 produced a 32.2 +/- 5.4% increase. Activated 
      pertussis toxin treatment of neutrophil plasma membranes inhibited by 72.5 +/- 
      14.3% the stimulation of GTPase activity induced by fMLP; however, activated 
      cholera toxin had no effect on the inhibition of fMLP stimulation, suggesting a 
      direct role for an Ni-like protein in the coupling process. In contrast to the 
      lack of inhibition of fMLP stimulation by activated cholera toxin treatment of 
      plasma membranes, both pertussis toxin and to a lesser extent cholera toxin 
      treatment reduced fMLP, C5a, and LTB4 stimulation of the GTPase in sonicates 
      prepared from pretreated whole cells. Pertussis toxin inhibited fMLP stimulation 
      of the GTPase by 75 +/- 7%, C5a stimulation was inhibited by 83 +/- 13%, and LTB4 
      stimulation was inhibited completely. Sonicates prepared from neutrophils treated 
      similarly with cholera toxin showed a smaller inhibition of GTPase activity (50 
      +/- 4% and 14 +/- 9% for fMLP and LTB4, respectively) with the exception of C5a, 
      where CT inhibition (81 +/- 32%) equaled pertussis toxin inhibition. Similarly, 
      pertussis toxin completely inhibited the release of the granule enzyme 
      N-acetyl-glucosaminidase by all three chemoattractants, whereas cholera toxin, 
      except with C5a stimulation, had little or no effect.(ABSTRACT TRUNCATED AT 400 
      WORDS)
FAU - Feltner, D E
AU  - Feltner DE
FAU - Smith, R H
AU  - Smith RH
FAU - Marasco, W A
AU  - Marasco WA
LA  - eng
GR  - HL-28445/HL/NHLBI NIH HHS/United States
PT  - Journal Article
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - J Immunol
JT  - Journal of immunology (Baltimore, Md. : 1950)
JID - 2985117R
RN  - 0 (Complement C5)
RN  - 0 (Ions)
RN  - 0 (Receptors, Complement)
RN  - 0 (Receptors, Formyl Peptide)
RN  - 0 (Receptors, Immunologic)
RN  - 0 (Virulence Factors, Bordetella)
RN  - 1HGW4DR56D (Leukotriene B4)
RN  - 59880-97-6 (N-Formylmethionine Leucyl-Phenylalanine)
RN  - 80295-54-1 (Complement C5a)
RN  - 9012-63-9 (Cholera Toxin)
RN  - EC 2.4.2.31 (Pertussis Toxin)
RN  - EC 3.1.3.2 (Phosphoric Monoester Hydrolases)
RN  - EC 3.2.1.- (Hexosaminidases)
RN  - EC 3.6.1.- (GTP Phosphohydrolases)
RN  - EC 3.6.1.- (GTP-Binding Proteins)
SB  - IM
MH  - Animals
MH  - Cell Membrane/enzymology
MH  - *Chemotaxis, Leukocyte
MH  - Cholera Toxin/pharmacology
MH  - Complement C5/physiology
MH  - Complement C5a
MH  - Enzyme Activation
MH  - GTP Phosphohydrolases/*physiology
MH  - GTP-Binding Proteins/*physiology
MH  - Hexosaminidases/metabolism
MH  - Ions
MH  - Leukotriene B4/physiology
MH  - N-Formylmethionine Leucyl-Phenylalanine/physiology
MH  - Neutrophils/*enzymology
MH  - Pertussis Toxin
MH  - Phosphoric Monoester Hydrolases/*physiology
MH  - Rabbits
MH  - Receptors, Complement/*physiology
MH  - Receptors, Formyl Peptide
MH  - Receptors, Immunologic/*physiology
MH  - Virulence Factors, Bordetella/pharmacology
EDAT- 1986/09/15 00:00
MHDA- 1986/09/15 00:01
CRDT- 1986/09/15 00:00
PHST- 1986/09/15 00:00 [pubmed]
PHST- 1986/09/15 00:01 [medline]
PHST- 1986/09/15 00:00 [entrez]
PST - ppublish
SO  - J Immunol. 1986 Sep 15;137(6):1961-70.