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G Schultz

Publications and source records attributed to G Schultz.

At least 217 records · Page 12Linked to original sources
PubMed1990

Activation of superoxide formation and lysozyme release in human neutrophils by the synthetic lipopeptide Pam3Cys-Ser-(Lys)4. Involvement of guanine-nucleotide-binding proteins and synergism with chemotactic peptides.

Upon exposure to the bacterial chemotactic peptide fMet-Leu-Phe, human neutrophils release lysozyme and generate superoxide anions (O2.-). The synthetic lipoamino acid N-palmitoyl-S-[2,3-bis(palmitoyloxy)-(2RS)-propyl]-(R)-cysteine (Pam3Cys), which is derived from the N-terminus of bacterial lipoprotein, when attached to Ser-(Lys)4 [giving Pam3Cys-Ser-(Lys)4], activated O2.- formation and lysozyme release in human neutrophils with an effectiveness amounting to about 15% of that of fMet-Leu-Phe. Palmitic acid, muramyl dipeptide, lipopolysaccharide and the lipopeptides Pam3Cys-Ala-Gly, Pam3Cys-Ser-Gly, Pam3Cys-Ser, Pam3Cys-OMe and Pam3Cys-OH did not activate O2.- formation. Pertussis toxin, which ADP-ribosylates guanine-nucleotide-binding proteins (G-proteins) and functionally uncouples formyl peptide receptors from G-proteins, prevented activation of O2.- formation by fMet-Leu-Phe and inhibited Pam3Cys-Ser-(Lys)4-induced O2.- formation by 85%. Lipopeptide-induced exocytosis was pertussis-toxin-insensitive. O2.- formation induced by Pam3Cys-Ser-(Lys)4 and fMet-Leu-Phe was enhanced by cytochalasin B, by a phorbol ester and by a diacylglycerol kinase inhibitor. Addition of activators of adenylate cyclase and removal of extracellular Ca2+ inhibited O2.- formation by fMet-Leu-Phe and Pam3Cys-Ser-(Lys)4 to different extents. Pam3Cys-Ser-(Lys)4 synergistically enhanced fMet-Leu-Phe-induced O2.- formation and primed neutrophils to respond to the chemotactic peptide at non-stimulatory concentrations. Our data suggest the following. (1) Pam3Cys-Ser-(Lys)4 activates neutrophils through G-proteins, involving pertussis-toxin-sensitive and -insensitive processes. (2) The signal transduction pathways activated by fMet-Leu-Phe and Pam3Cys-Ser-(Lys)4 are similar but not identical. (3) In inflammatory processes, bacterial lipoproteins and chemotactic peptides may interact synergistically to activate O2.- formation, leading to enhanced bactericidal activity.

Dose-Response Relationship, Drug↗
PubMed1990

A high-affinity bradykinin receptor in membranes from rat myometrium is coupled to pertussis toxin-sensitive G-proteins of the Gi family.

In rat myometrial membranes, two 3H-Bradykinin binding sites with KD values of 16 pM and 1.0 nM were identified. Employed at pM concentrations, bradykinin stimulated high affinity GTPases. This effect was abolished by the bradykinin antagonist, [D-Arg(Hyp3-Thi5,8, D-Phe7)]bradykinin (10 microM), and by treatment of membranes with pertussis toxin. Myometrial membranes contained two pertussis toxin substrates of 40 and 41 kDa, which corresponded immunologically to alpha-subunits of Gi-type G-proteins. The faster migrating substrate was tentatively identified as Gi2 alpha-subunit. The electrophoretic mobility of the slower migrating Gi alpha-subunit was very similar to that of the Gi3 alpha-subunit. Go alpha-subunits were not detected. Thus, in uterine smooth muscle, G-proteins of the Gi-family (Gi2, Gi3) couple high-affinity bradykinin receptors to their effector enzymes.

Adenosine Diphosphate Ribose↗
PubMed1990

Agonist-sensitive binding of a photoreactive GTP analog to a G-protein alpha-subunit in membranes of HL-60 cells.

Myeloid-differentiated HL-60 cells were used to study the activation of G-proteins by receptor agonists. Following incubation of membranes with the photoreactive GTP analog. [alpha-32P]GTP azidoanilide, and subsequent exposure to ultraviolet light (254 nm), photolabeling of 40 kDa proteins comigrating with the Gi2 alpha-subunit was observed. Photolabeling in the absence or presence of the chemoattractant, N-formyl-methionyl-leucyl-phenylalanine (FMLP), absolutely required Mg2+; FMLP stimulated photolabeling at all Mg2+ concentrations employed (up to 30 mM). Addition of GDP (3-50 microM) reduced basal photolabeling to a greater extent than photolabeling stimulated by FMLP. FMLP did not stimulate photolabeling of proteins modified by pertussis toxin. Leukotriene B4 and C5a also stimulated photolabeling of 40 kDa proteins. The results indicate that (i) the major G-protein in HL-60 cells, Gi2, requires Mg2+ for basal and receptor-stimulated activity, (ii) effective receptor-mediated activation of G-proteins is observed at mM concentrations of Mg2+, and (iii) receptor agonists apparently reduce the affinity of G-proteins for GDP.

Affinity Labels↗
PubMed1990

Calcium currents of neuroblastoma x glioma hybrid cells after cultivation with dibutyryl cyclic AMP and nickel.

The long-term modulation of calcium (Ca2+) currents (ICa) was studied in 108CC15 neuroblastoma x glioma hybrid (NxG) cells grown under various culture conditions. The following results were obtained: 1. Addition of 1 mM dibutyryl cyclic adenosine monophosphate (db-cAMP) or 0.1 microM forskolin to the culture medium increased a transient component of ICa two-fold within 3 days, from 21.0 +/- 1.6 pA/pF (n = 22) to a maximum of 40.0 +/- 2.6 pA/pF (n = 28). Under these conditions, cells also expressed a slowly inactivating ICa component (maximum after 3 days, 20.5 +/- 1.6 pA/pF, n = 28). 2. The fast inactivating ICa as well as the db-cAMP-induced slowly inactivating ICa were completely down-regulated during incubation of NxG cells with the inorganic Ca2+ channel blocker, nickel (Ni2+, 100 microM). The suppressing effect was reversed within 3 days of incubation in db-cAMP-containing medium lacking Ni2+. 3. Binding studies on membrane preparations of control and Ni2(+)-pretreated NxG cells revealed a marked difference in the maximal (+)3H-PN200-110 binding. The difference was seen in undifferentiated as well as in db-cAMP-incubated cells. 4. The protein synthesis blocker, cycloheximide, suppressed both the db-cAMP-induced increase and the reappearance of ICa following Ni2+ pretreatment. It is suggested that chronic application of db-cAMP or Ni2+ to NxG cells increases and decreases the number of Ca2+ channel proteins, respectively.

Animals↗
PubMed1990

Shikimate kinase from spinach chloroplasts : purification, characterization, and regulatory function in aromatic amino Acid biosynthesis.

Shikimate kinase was purified to near homogenity from spinach Spinacia oleracea L. chloroplasts and found to consist of a single 31 kilodalton polypeptide. The purified enzyme was unstable, but could be stabilized by a variety of added proteins, including oxidized and reduced thioredoxins. Whereas the isolated enzyme was stimulated by mono- and dithiol reagents, the enzyme in intact chloroplasts was unaffected by added thiols and showed only minor response to dark/light transitions. These results indicate that the previously reported stimulation of shikimate kinase activity by reduced thioredoxins is due to enzyme stabilization rather than to activation. In the current study, the purified enzyme was inhibited by added ADP and showed a strong response to energy charge. When intact chloroplasts were incubated in the dark in presence of shikimate, phosphoenolpyruvate and a source of ATP (dihydroxyacetone phosphate or ATP itself under appropriate conditions), aromatic amino acids were formed: phenylalanine and tyrosine. The data indicate that energy charge plays a role in regulating shikimate kinase, thereby controlling the shikimate pathway. An unidentified enzyme of the latter part of the pathway, leading from shikimate-3-phosphate to phenylalanine, appears to be activated by light.

Journal Article↗
PubMed1990

Plastidic Isoprenoid Synthesis during Chloroplast Development : Change from Metabolic Autonomy to a Division-of-Labor Stage.

The chloroplast isoprenoid synthesis of very young leaves is supplied by the plastidic CO(2) --> pyruvate --> acetyl-coenzyme A (C(3) --> C(2)) metabolism (D Schulze-Siebert, G Schultz [1987] Plant Physiol 84: 1233-1237) and occurs via the plastidic mevalonate pathway. The plastidic C(3) --> C(2) metabolism and/or plastidic mevalonate pathway of barley (Hordeum vulgare L.) seedlings changes from maximal activity at the leaf base (containing developing chloroplasts with incomplete thylakoid stacking but a considerable rate of photosynthetic CO(2)-fixation) almost to ineffectivity at the leaf tip (containing mature chloroplasts with maximal photosynthetic activity). The ability to import isopentenyl diphosphate from the extraplastidic space gradually increases to substitute for the loss of endogenous intermediate supply for chloroplast isoprenoid synthesis (change from autonomic to division-of-labor stage). Fatty acid synthesis from NaH(14)CO(3) decreases in the same manner as shown for leaf sections and chloroplasts isolated from these. Evidence has been obtained for a drastic decrease of pyruvate decarboxylase-dehydrogenase activity during chloroplast development compared with other anabolic chloroplast pathways (synthesis of aromatic amino acid and branched chain amino acids). The noncompetition of pyruvate and acetate in isotopic dilution studies indicates that both a pyruvate-derived and an acetate-derived compound are simultaneously needed to form introductory intermediates of the mevalonate pathway, presumably acetoacetyl-coenzyme A.

Journal Article↗
PubMed1990

New possibilities for platelet shape change evaluation using the laser rheoaggregometer.

The study of aggregation functions in blood platelets can at present be undertaken in vitro by measuring variations in intensity of transmitted light or electrical impedance change. These two methods allow only a general approach to the aggregation process. The laser rheoaggregometer was constructed to study early morphological modifications induced when the platelets are activated before aggregation. The results obtained are in keeping with the laws of light scattering and allow the measurement of platelet shape change.

Adenosine Diphosphate↗
PubMed1989

Phosphorylation of G-protein alpha-subunits in intact adipose cells: evidence against a mediating role in insulin-dependent metabolic effects.

The phosphorylation of G-protein alpha-subunits was studied in plasma membranes prepared from isolated, intact adipocytes equilibrated with [32P]phosphate and subsequently incubated in the presence or absence of insulin. In iodinated or unlabeled plasma membranes, antiserum generated against a peptide corresponding to a region common to G-protein alpha-subunits immunoprecipitated two major proteins of 45 and 40 kDa, which were identified as Gs and Gi alpha-subunit, respectively, by comparison with [32P]ADP-ribosylated G-proteins. In membranes prepared from cells equilibrated with [32P]phosphate, the antiserum precipitated a 45 kDa phosphoprotein. Pre-immune serum failed to immunoprecipitate the phosphoprotein. Insulin stimulated [32P]phosphate incorporation into the 45 kDa protein approximately 2-fold. Control experiments suggested that the 45 kDa phosphoprotein was not identical with G alpha s, since (1) the peptide used to raise the antiserum failed to inhibit significantly immunoprecipitation of the 45 kDa phosphoprotein with the antiserum, (2) in contrast to the Gs alpha-subunit, the phosphoprotein was readily removed from the immunocomplex by washing with sodium dodecyl sulfate (SDS), and (3) the subcellular localization of the phosphoprotein differed considerably from that of the Gs alpha-subunit. No phosphate was detected in immunoprecipitates from either basal or insulin-treated cells after the 45 kDa phosphoprotein had been removed. These data argue against a mediating role of phosphorylated G-protein alpha-subunits in the action of insulin.

Adipose Tissue↗
PubMed1989

Tissue distribution of beta 1- and beta 2-subunits of regulatory guanine nucleotide-binding proteins.

The beta-subunit of G-proteins occurs in two forms (beta 1 and beta 2), which differ in their primary structure as derived from cDNA clones and in their mobilities on SDS gels (36 and 35 kDa, respectively). To assess the tissue distribution of the two forms of beta-subunits, we synthesized peptides corresponding to defined regions of beta 1- and beta 2-subunits and injected them into rabbits; the antisera obtained reacted either with both beta-subunits or specifically with the beta 1- or the beta 2-subunit. They were used to identify the two beta-subunits in membranes prepared from various rat tissues and from human placenta. The concentration of total beta-subunits was high in rat brain and lung, human placenta, rat kidney, liver and spleen; it was much lower in rat erythrocytes, cardiac and skeletal muscle. In all tissues studied, both beta 1- and beta 2-subunits were detectable. In most tested tissues, the two forms were about equally distributed, whereas in the placenta, the beta 2-subunit was found to occur in approx. 2-fold excess over the beta 1-subunit. Our results demonstrate that both beta-subunits are widely distributed. In the majority of tissues, levels of beta 2-subunits are very similar to those of beta 1-subunits. Thus, the abundance of beta 2-subunits as compared to that of the beta 1-subunit is considerably higher than was previously estimated by measuring the respective mRNA levels.

Amino Acid Sequence↗
PubMed1989

The cytosolic G-protein alpha-subunit in human neutrophils responds to treatment with guanine nucleotides and magnesium.

The hydrodynamic properties of the Gi2 alpha-subunit in human neutrophil cytosol were analyzed. The S20,w value was 3.3 S at 30 degrees C and 4.1 S at 4 degrees C. Reconstitution of the cytosolic alpha-subunit with beta gamma-complex purified from porcine brain resulted in an S20,w value of 4.0 S at 30 degrees C and 4.2 S at 4 degrees C. Treatment of cytosol with G-protein-activating agents, GTP gamma S and MgCl2, decreased the S20,w value to 2.4 S at 30 degrees C and 2.9 S at 4 degrees C. Our results indicate that the cytosolic alpha-subunit of neutrophils is an inactive monomeric species at 30 degrees C capable of interacting with G-protein beta gamma-complex and responsive to treatment with activating agents leading to the two activated forms, alpha at 4 degrees C and alpha at 30 degrees C.

Animals↗
PubMed1989

Differential expression of cytosolic activation factors for NADPH oxidase in HL-60 leukemic cells.

Activation of NADPH oxidase in undifferentiated HL-60 leukemic cells and in HL-60 cells differentiated along the myeloid pathway with dibutyryl cyclic AMP (dbcAMP) or dimethyl sulfoxide (Me2SO) was studied. Upon stimulation with a calcium ionophore, a phorbol ester, arachidonic acid or gamma-hexachlorocyclohexane, Me2SO-differentiated HL-60 cells generated superoxide (O2-) at higher rates than dbcAMP-differentiated cells. Undifferentiated cells generated O2- only at low rates upon stimulation with the above agents. In cell-free systems, NADPH oxidase activity was reconstituted by combining membranes of undifferentiated or dbcAMP- or Me2SO-differentiated HL-60 cells, cytosol of Me2SO-differentiated cells and arachidonic acid. This basal O2- formation was enhanced several-fold by guanosine 5'-O-(3-thiotriphosphate) (GTP[gamma S]), a potent activator of guanine nucleotide-binding proteins. In contrast, cytosol of dbcAMP-differentiated cells reconstituted O2- formation only in the presence of GTP[gamma S], and cytosol of undifferentiated cells was inactive. Submaximally stimulatory amounts of cytosolic protein of Me2SO- and dbcAMP-differentiated cells synergistically stimulated O2- formation in the presence but not in the absence of GTP[gamma S]. We conclude that differentiations of HL-60 cells with Me2SO and dbcAMP are not equivalent with respect to activation of NADPH oxidase and that two cytosolic activation factors are involved in the regulation of this effector system.

Arachidonic Acid↗
PubMed1989

Activation of NADPH oxidase by purine and pyrimidine nucleotides involves G proteins and is potentiated by chemotactic peptides.

Human neutrophils and HL-60 leukaemic cells possess an NADPH oxidase which catalyses superoxide (O2-) formation and is activated by the chemotactic peptide, N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMet-Leu-Phe). In dibutyryl cyclic AMP-differentiated HL-60 cells, ATP and UTP in the presence of cytochalasin B activated O2- formation with EC50 values of 5 microM and efficacies amounting to 30% of that of fMet-Leu-Phe. The potency order of purine nucleotides in activating O2- generation was ATP = adenosine 5'-O-(3-thiotriphosphate) greater than ITP greater than dATP = ADP. Pyrimidine nucleotides activated NADPH oxidase in the potency order UTP greater than dUTP greater than CTP = TTP = UDP. Pertussis toxin completely prevented activation of NADPH oxidase by fMet-Leu-Phe and UTP, whereas the effect of ATP was only partially inhibited. ATP and UTP enhanced O2- generation induced by fMet-Leu-Phe by up to 8-fold, and primed the cells to respond to non-stimulatory concentrations of fMet-Leu-Phe. Activation of NADPH oxidase by UTP but not by ATP was inhibited by various activators of adenylate cyclase. In dimethyl sulphoxide-differentiated HL-60 cells and in human neutrophils, ATP and UTP per se did not activate NADPH oxidase, but they potentiated the effect of fMet-Leu-Phe. Our results suggest that purine and pyrimidine nucleotides act via purino- and novel pyrimidinoceptors respectively, which are coupled to guanine nucleotide-binding proteins leading to the activation of NADPH oxidase. As ATP and UTP are released from cells under physiological and pathological conditions, these nucleotides may play roles as intercellular signal molecules in the activation of O2- formation.

Adenosine Triphosphate↗
PubMed1989

Purine and pyrimidine nucleotides potentiate activation of NADPH oxidase and degranulation by chemotactic peptides and induce aggregation of human neutrophils via G proteins.

Whereas the chemotactic peptide, N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMet-Leu-Phe), induced NADPH-oxidase-catalyzed superoxide (O2-) formation in human neutrophils, purine and pyrimidine nucleotides per se did not stimulate NADPH oxidase but enhanced O2- formation induced by submaximally and maximally stimulatory concentrations of fMet-Leu-Phe up to fivefold. On the other hand, FMet-Leu-Phe primed neutrophils to generate O2- upon exposure to nucleotides. At a concentration of 100 microM, purine nucleotides enhanced O2- formation in the effectiveness order adenosine 5'-O-[3-thio]triphosphate (ATP[gamma S]) greater than ITP greater than guanosine 5'-O-[3-thio]triphosphate (GTP[gamma S]) greater than ATP = adenosine 5'-O-[2-thio]triphosphate (Sp-diastereomer) = GTP = guanosine 5'-O-[2-thio]diphosphate (GDP[beta S] = ADP greater than adenosine 5'-[beta, gamma-imido]triphosphate = adenosine 5'-O-[2-thio]triphosphate] (Rp-diastereomer). Pyrimidine nucleotides stimulated fMet-Leu-Phe-induced O2- formation in the effectiveness order uridine 5'-O-[3-thio]triphosphate (UTP[gamma S]) = UTP greater than CTP. Uracil (UDP[beta S]) = uridine 5'-O[2-thio]triphosphate (Rp-diastereomer) (Rp)-UTP[beta S]) = UTP greater than CTP. Uracil nucleotides were similarly effective potentiators of O2- formation as the corresponding adenine nucleotides. GDP[beta S] and UDP[beta S] synergistically enhanced the stimulatory effects of ATP[gamma S], GTP[gamma S] and UTP[gamma S]. Purine and pyrimidine nucleotides did not induce degranulation in neutrophils but potentiated fMet-Leu-Phe-induced release of beta-glucuronidase with similar nucleotide specificities as for O2- formation. In contrast, nucleotides per se induced aggregation of neutrophils. Treatment with pertussis toxin prevented aggregation induced by both nucleotides and fMet-Leu-Phe. Our results suggest that purine and pyrimidine nucleotides act via nucleotide receptors, the nucleotide specificity of which is different from nucleotide receptors in other cell types. Neutrophil nucleotide receptors are coupled to guanine-nucleotide-binding proteins. As nucleotides are released from cells under physiological and pathological conditions, they may play roles as intercellular signal molecules in neutrophil activation.

Cell Aggregation↗
PubMed1989

Secretion-stimulating and secretion-inhibiting hormones stimulate high-affinity pertussis-toxin-sensitive GTPases in membranes of a pituitary cell line.

Different peptide hormones influence hormone secretion in pituitary cells by diverse second messenger systems. Recent data indicate that luteinizing-hormone-releasing hormone (LHRH) stimulates and somatostatin inhibits voltage-dependent Ca2+ channels of GH3 cells via pertussis-toxin-sensitive mechanisms [Rosenthal et al. (1988) EMBO J. 7, 1627-1633]. In other pituitary cell lines, somatostatin has been shown to cause a pertussis-toxin-sensitive decrease in adenylate cyclase activity, and LHRH and thyrotropin-releasing hormone (TRH) stimulate phosphoinositol lipid hydrolysis in a pertussis-toxin-independent manner. Whether stimulation of Ca2+ influx by TRH is affected by pertussis toxin is not known. In order to elucidate which of the hormone receptors interact with pertussis-toxin-sensitive and -insensitive G-proteins, we measured the effects of LHRH, somatostatin and TRH on high-affinity GTPases in membranes of GH3 cells. In control membranes, both LHRH and TRH stimulated the high-affinity GTPase by 20%, somatostatin by 25%. Maximal hormone effects were observed at a concentration of about 1 microM. Pretreatment of cells with pertussis toxin abolished pertussis-toxin-catalyzed [32P]ADP-ribosylation of 39-40-kDa proteins in subsequently prepared membranes and reduced basal GTPase activity. The toxin also reduced by more than half the increases in GTPase activity induced by LHRH and TRH; stimulation of GTPase by somatostatin was completely suppressed. Stimulation of adenylate cyclase by vasoactive intestinal peptide (VIP) was not impaired by pretreatment of cells with pertussis toxin. Somatostatin but not LHRH and TRH decreased forskolin-stimulated adenylate cyclase activity. The results suggest that the activated receptors for LHRH and TRH act via pertussis-toxin-sensitive and -insensitive G-proteins, whereas effects of somatostatin are exclusively mediated by pertussis-toxin-sensitive G-proteins.

Adenylate Cyclase Toxin↗
PubMed1989

Lack of effect of opioid peptides, morphine and naloxone on superoxide formation in human neutrophils and HL-60 leukemic cells.

There are controversial reports in the literature concerning the effects of opioids on superoxide (O2-) formation in phagocytes, these agents being either inhibitory or stimulatory. We re-examined this issue and compared the effects of the chemotactic peptide, N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMet-Leu-Phe), phorbol myristate acetate (PMA), ATP, platelet activating factor (PAF), cytochalasin B (CB) and prostaglandin E1 (PGE1) with those of various opioids on O2- formation in human neutrophils and HL-60 leukemic cells under defined experimental conditions. In the presence of CB, fMet-Leu-Phe and PAF concentration-dependently activated O2- formation in neutrophils with EC50 values of 20 nM and 100 nM, respectively. In the absence of CB, fMet-Leu-Phe and PAF were much less effective. PAF synergistically enhanced O2- formation induced by fMet-Leu-Phe. ATP at a concentration of 100 microM and the opioids, methionine enkephalin, beta-endorphin, dynorphin, [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin, [D-Ala2-D-Leu5]-enkephalin and morphine at concentrations between 10 pM to 1 microM did not activate O2- formation. ATP but not beta-endorphin potentiated fMet-Leu-Phe-induced O2- formation. O2- formation induced by a maximally stimulatory concentration of PMA (100 ng/ml) was enhanced by fMet-Leu-Phe but was unaffected by methionine enkephalin or PGE1. PMA at a non-stimulatory concentration (2 ng/ml) potentiated the effect of fMet-Leu-Phe but did not induce responsiveness to PAF, ATP or beta-endorphin. PGE1 strongly inhibited fMet-Leu-Phe-induced O2- formation, whereas morphine, methionine enkephalin and the opioid antagonist, naloxone, were without effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1989

Export and activity of hybrid FhuA'-'Iut receptor proteins and of truncated FhuA' proteins of the outer membrane of Escherichia coli.

The FhuA protein in the outer membrane of Escherichia coli serves as a multifunctional receptor for the phages T5, T1, phi 80, for colicin M, for ferrichrome (Fe3+-siderophore) and for the structurally related antibiotic, albomycin. To determine structural domains required for these receptor functions and for export, a fusion protein between FhuA and Iut (receptor for Fe3+-aerobactin and cloacin DF13) was constructed. In the FhuA'-'Iut hybrid protein, 24 amino acids of FhuA were replaced by 19 amino acids, 18 of which were from Iut. The number of plaque forming units of phage T5 and T1 on cells expressing FhuA'-'Iut was nearly as high as on cells expressing plasmid-encoded wild-type FhuA. However, 10(7)-fold higher concentrations of phage phi 80 and 10(3) times more colicin M were required to obtain a zone of growth inhibition. Truncated FhuA' proteins in which the last 24 amino acids at the carboxy-terminus were replaced by 16 (FhuA'2) or 3 (FhuA'T) amino acids could hardly be detected on polyacrylamide electrophoretograms of outer membrane proteins, due to proteolytic degradation. Sensitivity of cells expressing FhuA'2 to phage T5 and T1 was reduced by several orders of magnitude and sensitivity to phage phi 80 and colicin M was totally abolished. In contrast, cells expressing FhuA'T were nearly as sensitive to pahge T5, T1, and phi 80 and to colicin M as cells containing FhuA'-'Iut. None of the constructs could grow on ferrichrome as sole iron source and none was sensitive to albomycin.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗