PubMed Health⌕ Search

Biomedical subjects

F J Walker

Publications and source records attributed to F J Walker.

At least 19 recordsLinked to original sources
PubMed2003

The interface phase and the Schottky barrier for a crystalline dielectric on silicon.

The barrier height for electron exchange at a dielectric-semiconductor interface has long been interpreted in terms of Schottky's theory with modifications from gap states induced in the semiconductor by the bulk termination. Rather, we show with the structure specifics of heteroepitaxy that the electrostatic boundary conditions can be set in a distinct interface phase that acts as a "Coulomb buffer." This Coulomb buffer is tunable and will functionalize the barrier-height concept itself.

Journal Article↗
PubMed2001

Physical structure and inversion charge at a semiconductor interface with a crystalline oxide.

We show that the physical and electrical structure and hence the inversion charge for crystalline oxides on semiconductors can be understood and systematically manipulated at the atomic level. Heterojunction band offset and alignment are adjusted by atomic-level structural and chemical changes, resulting in the demonstration of an electrical interface between a polar oxide and a semiconductor free of interface charge. In a broader sense, we take the metal oxide semiconductor device to a new and prominent position in the solid-state electronics timeline. It can now be extensively developed using an entirely new physical system: the crystalline oxides-on-semiconductors interface.

Journal Article↗
PubMed2000

Influenza, pneumococcal, and tetanus toxioid vaccination of adults--United States, 1993-7.

PROBLEM/CONDITION: An increasing proportion of adults have received recommended vaccinations against influenza, pneumococcal infection, and tetanus. However, in 1995, fewer than 60% of adults were vaccinated as recommended. REPORTING PERIOD COVERED: 1993-1997. DESCRIPTION OF SYSTEM: Data were obtained from the state-based Behavioral Risk Factor Surveillance System (BRFSS) for 1993, 1995, and 1997 and from the National Health Interview Survey (NHIS) for 1995 to describe national, regional, and state-specific patterns of use of influenza and pneumococcal vaccines and tetanus toxoid among noninstitutionalized adults aged > or = 18 years. RESULTS: Among adults aged > or = 65 years in 1995, 58% reported receiving an influenza vaccination during the previous 12 months, and 34% reported ever receiving a pneumococcal vaccination. In this age group, non-Hispanic whites were more likely to report receipt of influenza (61%) and pneumococcal vaccines (36%) than non-Hispanic blacks (40% and 22%, respectively) and Hispanics (50% and 23%, respectively). Among the 50 states and the District of Columbia, the median vaccination level among older adults (i.e., persons aged > or = 65 years) increased from 51% in 1993 to 66% in 1997 for influenza vaccine, and from 28% in 1993 to 46% in 1997 for pneumococcal vaccine. Adults with chronic medical conditions had low vaccination levels. Those aged 50-64 years were more likely than those aged 18-49 years to report influenza (38% versus 20%) and pneumococcal vaccination (20% versus 12%). In 1995, the proportion of adults who reported receiving a tetanus vaccination during the previous 10 years decreased with age, from 65% among those aged 18-49 years to 54% among those aged 50-64 years and to 40% among those aged > or = 65 years. In each age group, women were less likely than men to report receiving tetanus toxoid; and among adults aged > or = 65 years, Hispanics and Asians/Pacific Islanders were least likely among all racial/ethnic groups to report receiving tetanus toxoid. INTERPRETATION: By 1995, the Healthy People 2000 objective to increase to at least 60% the proportion of persons aged > or = 65 years who had received annual influenza vaccination had been achieved among non-Hispanic whites (objective 20.11). However, substantial improvement is needed among non-Hispanic blacks, Hispanics, and adults aged < 65 years with high-risk medical conditions. PUBLIC HEALTH ACTIONS: Continued surveillance of vaccine coverage among adults will direct attention to undervaccinated populations that may be disproportionately affected by vaccine-preventable diseases. Vaccination coverage data can be used to guide efforts to increase awareness among health-care providers and the public about the benefits of vaccination, establish systems to ensure that every contact with the health-care system is used to update vaccinations, and further support financial mechanisms to increase vaccine delivery.

Adult↗
PubMed1996

Activated protein C-catalyzed proteolysis of factor VIIIa alters its interactions within factor Xase.

Factor VIIIa, the cofactor for the factor IXa-dependent conversion of factor X to factor Xa, is proteolytically inactivated by activated protein C (APC). APC cleaves at two sites in factor VIIIa, Arg336, near the C terminus of the A1 subunit; and Arg562, bisecting the A2 subunit (Fay, P., Smudzin, T., and Walker, F. (1991) J. Biol. Chem. 266, 20139-20145). Factor VIIIa increased the fluorescence anisotropy of fluorescein-Phe-Phe-Arg factor IXa (Fl-FFR-FIXa; Kd = 42.4 nM), whereas cleavage of factor VIIIa by APC eliminated this property. Isolation of the APC-cleaved A1/A3-C1-C2 dimer (A1336/A3-C1-C2), and the fragments derived from cleaved A2 subunit (A2N/A2C), permitted dissection of the roles of individual cleavages in cofactor inactivation. Intact A1/A3-C1-C2 dimer increased Fl-FFR-FIXa anisotropy and bound factor X in a solid phase assay, while these activities were absent in the A1336/A3-C1-C2. However, the residues removed by this cleavage, Met337 Arg372, did not directly participate in these functions since neither a synthetic peptide to this sequence nor an anti-peptide polyclonal antibody blocked these activities using intact dimer. CD spectral analysis of the intact and truncated dimers indicated reduced alpha and/or beta content in the latter. The A1/A3-C1-C2 dimer plus A2 subunit reconstitutes cofactor activity and produced a factor VIIIa-like effect on the anisotropy of Fl-FFR-FIXa. However, when A2 was replaced by the A2N/A2C fragments, the resulting fluorescence signal was equivalent to that observed with the dimer alone. These results indicate that APC inactivates the cofactor at two levels within the intrinsic factor Xase complex. Cleavage of either subunit modulates the factor IXa active site, suggesting an essential synergy of interactive sites in factor VIIIa. Furthermore, cleavage of the A1 site alters the conformation of a factor X binding site within that subunit, thereby reducing the affinity of cofactor for substrate.

Amino Acid Sequence↗
PubMed1994

Factor IXa protects factor VIIIa from activated protein C. Factor IXa inhibits activated protein C-catalyzed cleavage of factor VIIIa at Arg562.

Factor VIIIa is inactivated by both factor IXa and activated protein C. The latter protease rapidly attacked a site at Arg562 (A2 subunit), whereas both proteases slowly cleaved factor VIIIa at Arg336 (A1 subunit). Cofactor inactivation catalyzed by activated protein C was 8-fold faster than that catalyzed by factor IXa. Simultaneous reaction of factor VIIIa with the two enzymes resulted in a rate of inactivation intermediate to that observed for the individual proteases. Under these conditions, the activated protein C-catalyzed cleavage at Arg562 was inhibited such that cofactor inactivation resulted primarily from cleavage at Arg336. Substitution of factor IXa modified in its active site with 6-(dimethylamino)-2-naphthalenesulfonyl-glutamylglycylarginyl++ + chloromethyl ketone (DEGR-IXa) for the native enzyme yielded a similar rate of activated protein C-catalyzed cleavage at the A1 site, whereas cleavage at the A2 site was virtually eliminated. However, the inclusion of protein S resulted in a marked increase in cleavage at the A2 site that correlated with an increased rate of cofactor inactivation. Active site-modified activated protein C inhibited the factor IXa-dependent enhancement of factor VIIIa reconstitution from isolated subunits. In addition, the factor VIIIa-dependent fluorescence enhancement of DEGR-activated protein C was inhibited by EGR-IXa. These results indicate that factor IXa can reduce the rate of activated protein C-catalyzed cofactor inactivation by selectively blocking cleavage at the A2 domainal site, an effect reversed by protein S. One mechanism consistent with the reciprocal inhibitory effects of the proteases is that activated protein C and factor IXa occupy overlapping sites on the cofactor. Thus, factor IXa may protect factor VIIIa by preventing activated protein C binding.

Amino Acid Chloromethyl Ketones↗
PubMed1992

Characterization of the interaction between the heavy and light chains of bovine factor Va.

Bovine factor Va has been previously been shown to consist of heavy (M(r) = 94,000) and light chains (M(r) = 81,000), that interact in a manner dependent upon the presence of either calcium or manganese ions. In an attempt to understand the mechanism of subunit interaction we have studied the effects of temperature and ions on factor Va stability. The rates of formation of factor Va from isolated chains and dissociation were temperature-dependent with an energy of activation of 6.2 and 1.3 kcal mol-1, respectively. The yield of factor Va from isolated chains was inversely related to the amount of time the chains were incubated at 4 degrees C. Incubation of individual chains revealed that the heavy chain is cold-labile, an effect that is reversible. Manganese ion was observed to prevent the conversion to the inactive form. High salt tends to stabilize the two-chain structure of factor Va, but is inhibitory to its formation from isolated chains. High concentrations of either manganese or calcium ions also inhibited reconstitution of activity. The light chain, in particular, was sensitive to the presence of manganese or calcium ion. Heavy chain that had been cleaved by activated protein C had a weakened interaction with the light chain, and the resulting complex had no procoagulant activity. Cooling of the heavy chain to 4 degrees C enhanced its intrinsic fluorescence. Manganese ion prevented some of this enhancement. The heavy chain fluorescence returned to the room temperature value with a half-life of approximately 10 min. In the presence of manganese ion relaxation was accelerated. The intrinsic fluorescence of activated protein C-cleaved heavy chain was not increased when the temperature was decreased. These data suggest that the heavy chain can exist in two forms. Elevated temperature converts it to a form that can bind ions and have a productive interaction with the light chain. However, conditions that prevent the heavy chain from combining with the light chain also stabilize the two subunit structure, suggesting that the high affinity of the complex is due to conformational changes that occur after chain interaction.

Animals↗
PubMed1992

The thrombotic diathesis associated with the presence of phospholipid antibodies may be due to low levels of free protein S.

PURPOSE: To determine if abnormalities in the protein C/protein S anticoagulant system exist in patients with phospholipid antibodies who had the primary clinical complaint of fetal wastage. PATIENTS AND METHODS: Eleven patients with fetal wastage and phospholipid antibodies were selected for study. Some patients also gave a history of previous thrombotic events related to oral contraceptives and/or pregnancy, but patients were not selected because of a history of clinical thrombosis. The levels of protein C (chromogenic assay), protein S (both free and bound) (Laurell rocket), and C4b-binding protein (Laurell rocket) were measured, and assays for the presence of antibodies against protein S or protein C were performed. RESULTS: Seven of the 11 patients were found to have low levels of free protein S. Total protein S and protein C levels were within the normal range in all patients. Antibodies to protein C and protein S were not found in any patient. These findings suggest that free protein S levels may be abnormally low in some patients with phospholipid antibodies. CONCLUSION: Free protein S levels are abnormally low in some patients with phospholipid antibodies, and this abnormality may be a factor contributing to the thrombotic diathesis associated with phospholipid antibodies.

Abortion, Habitual↗
PubMed1992

Regulation of blood coagulation by the protein C system.

Protein C is a plasma, vitamin K-dependent zymogen of a serine protease that can inhibit blood coagulation. Protein C is regulated by a series of reactions known as the protein C pathway. The importance of this pathway is seen in the occurrence of thrombosis in individuals with deficiencies in elements of the pathway like protein C and protein S. Work on several steps in this pathway has revealed that mechanisms involved in activation of protein C and the expression of its anticoagulant activity have features that allow for the expression of the anticoagulant activity away from sites in which procoagulant reactions occur, but not systemically. Thrombin, the principal procoagulant enzyme at the site of an injury, is converted to an anticoagulant enzyme at distant sites through its interaction with the endothelial cell protein thrombomodulin. Structural and functional studies have revealed the importance of several domain structures in the modulation of thrombin activity. Structural features of both activated protein C and its substrates (coagulation factors V and VIII) are such that they require the localization of enzyme and substrate on the surface of phosphatidyl serine containing membranes for optimum activity.

Animals↗
PubMed1991

Activated protein C-catalyzed inactivation of human factor VIII and factor VIIIa. Identification of cleavage sites and correlation of proteolysis with cofactor activity.

Human factor VIII and factor VIIIa were proteolytically inactivated by activated protein C. Cleavages occurred within the heavy chain (contiguous A1-A2-B domains) of factor VIII and in the heavy chain-derived A1 and A2 subunits of factor VIIIa, whereas no proteolysis was observed in the light chain or light chain-derived A3-C1-C2 subunit. Reactivity to an anti-A2 domain monoclonal antibody and NH2-terminal sequence analysis of three terminal digest fragments from factor VIII allowed ordering of fragments and identification of cleavage sites. Fragment A1 was derived from the NH2 terminus and resulted from cleavage at Arg336-Met337. The A2 domain was bisected following cleavage at Arg562-Gly563 and yielded fragments designated A2N and A2C. A third cleavage site is proposed at the A2-B junction (Arg740-Ser741) since fragment A2C was of equivalent size when derived either from factor VIII or factor VIIIa. The site at Arg562 was preferentially cleaved first in factor VIII(alpha) compared with the site at Arg336, and it was this initial cleavage that most closely correlated with the loss of cofactor activity. Factor VIIIa was inactivated 5-fold faster than factor VIII, possibly as a result of increased protease utilization of the site at Arg562 when the A2 subunit is not contiguous with the A1 domain. When initial cleavage occurred at Arg336, it appeared to preclude subsequent cleavage at Arg562, possibly by promoting dissociation of the A2 domain (subunit) from the A1/light chain dimer. This conclusion was supported by the failure of protease treated A1/A3-C1-C2 dimer to bind A2 subunit and gel filtration analysis that showed dissociation of the A2 domain-derived fragments, A2N and A2C, from the A1 fragment/light chain dimer. These results suggest a mechanism for activated protein C-catalyzed inactivation of factor VIII(alpha) involving both covalent alteration and fragment dissociation.

Amino Acid Sequence↗
PubMed1991

Species specificity of the fibrinolytic effects of activated protein C.

Activated protein C (APC) has been shown to stimulate fibrinolysis in both in vitro and in vivo experimental systems. In order to test the importance of protein S in the fibrinolytic activity of APC we have compared the activity of APC, prepared from rabbit, bovine and human plasma, in the stimulation of whole blood clot lysis, the inactivation of plasminogen activator inhibitors and anticoagulant activity. When whole blood clot lysis was performed using tissue plasminogen activator in either human or rabbit blood, APC was found to enhance clot lysis in a species specific manner that paralleled the pattern observed for its anticoagulant activity. Bovine APC, was the poorest stimulator of fibrinolysis in human plasma. However, if bovine protein S was also added to human plasma, bovine APC was as effective in promoting fibrinolysis as human APC. In contrast, no species specific effects on the inactivation of plasminogen activator inhibitor activity was observed. Though substantial effects of APC on plasminogen activator inhibitor levels were made by rabbit, human and bovine activated protein C in human plasma, there was no effect of activated protein C on the rate of clot lysis of human plasma. These results suggest that protein S is important for the expression of the fibrinolytic activity of activated protein C and that the effect of protein S may be useful for the differentiation of fibrinolytic effects of activated protein C that may be related to the inactivation of plasminogen activator inhibitors and those that are not.

Animals↗
PubMed1991

von Willebrand factor mediates protection of factor VIII from activated protein C-catalyzed inactivation.

Factor VIII, a cofactor of the intrinsic clotting pathway, is proteolytically inactivated by the vitamin K-dependent serine protease, activated protein C in a reaction requiring Ca2+ and a phospholipid surface. Factor VIII was inactivated 15 times faster than factor VIII in complex with either von Willebrand factor (vWf) or the large homodimeric fragment, SPIII (vWf residues 1-1365). Free factor VIII or factor VIII in complex with a smaller fragment, SPIII-T4 (vWf residues 1-272), were inactivated at the same rate, suggesting that this effect was dependent upon the size of factor VIII-vWf complex rather than changes in factor VIII brought about by occupancy of the vWf-binding site. Thrombin cleavage of the factor VIII light chain to remove the vWf-binding site eliminated the protective effects of vWf. In the absence of phospholipid, high levels of the protease inactivated both free and vWf-bound factor VIII at equivalent rates. Using the same conditions, isolated heavy chains and the heavy chains of factor VIII were proteolyzed at similar rates. Taken together, these results suggested that, in the absence of phospholipid, inactivation of factor VIII is independent of factor VIII light chain and further suggest that vWf did not mask susceptible cleavage sites in the cofactor. Solution studies employing fluorescence energy transfer using coumarin-labeled factor VIII (fluorescence donor) and synthetic phospholipid vesicles labeled with octadecyl rhodamine (fluorescence acceptor) indicated saturable binding and equivalent extents of donor fluorescence quenching for factor VIII alone or when complexed with SPIII-T4. However, complexing of factor VIII with either vWf or SPIII eliminated its binding to the phospholipid. Since a phospholipid surface is required for efficient catalysis by the protease, these results suggest that vWf protects factor VIII by inhibiting cofactor-phospholipid interactions.

Electrophoresis, Polyacrylamide Gel↗
PubMed1990

Purification and preliminary characterization of rabbit vitamin K-dependent coagulation proteins.

We describe herein a procedure for the purification of protein C, protein S, prothrombin, factor VII, and factor X to apparent homogeneity from rabbit plasma. The initial steps, which are common to the purification of vitamin K-dependent proteins from other mammalian species, include adsorption onto and elution from barium followed by anion exchange chromatography. Proteins were further purified using a variety of techniques, including affinity chromatography, gel filtration, and anion exchange chromatography in a Fast Protein Liquid Chromatography system. Significant structural homologies exist between rabbit, human, and bovine vitamin K-dependent proteins. As is true for protein C and factor X in human and bovine plasma, rabbit protein C and factor X are two-chain proteins which can be converted to active proteases by specific venom activators. Rabbit factor VII is also a two-chain protein and can restore coagulant activity to human or bovine plasma deficient in factor VII. In contrast, rabbit protein S and prothrombin are single chain proteins. In view of the well-described species specificity of many of the vitamin K-dependent proteins, purified rabbit coagulant and anticoagulant proteins should be useful in the development of animal models of coagulation and/or thrombosis.

Animals↗
PubMed1990

Characterization of an interaction between protein C and ceruloplasmin.

Coagulation factors V and VIII are substrates for activated protein C. Binding sites for the protease have been localized to homologous sequences within the terminal A domains of these proteins. Since ceruloplasmin contains significant sequence homology to these domains, a study was undertaken to determine whether ceruloplasmin was an activated protein C-binding protein. Ceruloplasmin was observed to inhibit the activated protein C-catalyzed inactivation of both factor Va and factor VIII. Searches of the ceruloplasmin sequence revealed a decapeptide sequence, HAGMETTYTV (residues 1028-1037) that shares 60 and 40% sequence identity with the activated protein C binding sequence in factors VIII and V, respectively. This peptide also inhibited factor Va inactivation and in addition was observed to enhance the amidolytic activity of activated protein C. The ferrous oxidase activity of ceruloplasmin was stimulated 5-fold by activated protein C, and this effect was negated by the peptide HAGMETTYTV. These results indicate that these conserved sequences of ceruloplasmin and factors V and VIII interact with activated protein C and suggest that this region may be important in the regulation of this anticoagulant protein.

Amino Acid Sequence↗
PubMed1990

Identification of the binding site for activated protein C on the light chain of factors V and VIII.

Activated protein C has been observed to bind to the light chains of factor Va and factor VIII. Fragments of the factor VIII light chain were produced by recombinant DNA techniques and expressed in Escherichia coli. Three fragments of the light chain were studied; L4 (residues 1974-2332), L3.2 (residues 1560-1829 and 2046-2332), and L3.3 (residues 1560-2052). Two fragments, L4 and L3.3, which overlapped sequences between residues 1974-2052, inhibited the anticoagulant activity of activated protein C. Comparison of the sequences of factors V and VIII in this region revealed that residues 2005-2018 in the factor VIII sequence were homologous with residues 1861-1874 in the factor V sequence. The peptides Arg-Ala-Gly-Met-Gln-Thr-Phe-Leu-Ile (RAGMQTPFLI; residues 1865-1874) from the factor V sequence and His-Ala-Gly-Met-Ser-Thr-Leu-Phe-Ile-Val (HAGMSTLFIV; residues 2009-2018) from the factor VIII sequence were synthesized. Both peptides were observed to inhibit the anticoagulant activity of activated protein C and its inactivation of factors Va and VIII. Furthermore RAGMQTPFLI quenched the fluorescence of the dansyl-Glu-Gly-Arg-modified protease. Polyclonal antibodies against RAGMQTPFLI bound to factor Va and inhibited the anticoagulant activity of activated protein C and the inactivation of factor Va. These results indicate that a portion of the binding sites for activated protein C on the light chains of factors V and VIII are contained in the sequences RAGMQTPFLI or HAGMSTLFIV, respectively.

Amino Acid Sequence↗
PubMed1990

Enhancement of rabbit protein S anticoagulant cofactor activity in vivo by modulation of the protein S C4B binding protein interaction.

The carboxy-terminal region of protein S has been recently been observed to be involved in the interaction between protein S and C4b-binding protein (Walker, F. J. 1989. J. Biol. Chem. 264:17645-17658). A synthetic peptide, GVQLDLDEAI, corresponding to that region of protein S has been used to investigate the protein S/C4b-binding protein interaction in vitro and in vivo. Rabbit activated protein C possesses species-specific anticoagulant activity for which rabbit protein S functions as a cofactor. In plasma, rabbit protein S is found in complex with C4b-binding protein. GVQLDLDEAI can inhibit this interaction, resulting in enhancement of the anticoagulant activity of rabbit activated protein C. The effect of the peptide can be blocked by the concurrent addition of human or rabbit C4b-binding protein. When infused into rabbits, GVQLDLDEAI was cleared from the circulation with a half-life of 80 min. This is significantly less rapid than the clearance of similarly sized control peptides (half-life of 15 min), but much more than that of bovine protein S, a much larger protein (half-life of 15 h). Plasma samples removed from the rabbits after infusion with GVQLDLDEAI were found to have increased concentrations of free protein S and to show enhanced anticoagulation by rabbit activated protein C ex vivo in a dose-dependent manner. The concentration for half-maximal effect (5 microM) was very similar to that observed in vitro. These results suggest that the formation of a complex between protein S and C4b-binding protein is important in the regulation of protein S activity in vivo, and that modulation of this interaction allows one to influence the anticoagulant activity of the protein C pathway.

Amino Acid Sequence↗