The Breathing Mode in Extended Skyrme Model

We study an extended Skyrme model which includes fourth and sixth-order terms. We explore some static properties like the $\Delta$-nucleon mass splitting and investigate the Skyrmion breathing mode in the framework of the linear response theory. We find that the monopole response function has a pronounced peak located at $\sim$ 400 MeV, which we identify to the Roper resonance $N(1440)$. As compared to the standard one, the extended Skyrme model provides a more accurate description of baryon properties.Comment: 12 pages of plain Latex and 3 figures (available from the authors), preprint IPNO/TH 93-0

Effective chiral Lagrangians for spin-1 mesons

The commonly used types of effective theory for vector mesons are reviewed and their relationships clarified. They are shown to correspond to different choices of field for spin-1 particles and the rules for transforming between them are described. The importance of respecting chiral symmetry is stressed. The choice of fields that transform homogeneously under the nonlinear realisation of chiral symmetry imposes no preconceptions about the types of coupling for the mesons. This representation thus provides a convenient framework for relating different theories. It is also used to elucidate the nature of the assumptions in specific hidden-gauge and massive Yang-Mills models that have been widely used.Comment: 46 pages (RevTeX

On different lagrangian formalisms for vector resonances within chiral perturbation theory

We study the relation of vector Proca field formalism and antisymmetric tensor field formalism for spin-one resonances in the context of the large N_C inspired chiral resonance Lagrangian systematically up to the order O(p6) and give a transparent prescription for the transition from vector to antisymmetric tensor Lagrangian and vice versa. We also discuss the possibility to describe the spin-one resonances using an alternative "mixed" first order formalism, which includes both types of fields simultaneously, and compare this one with the former two. We also briefly comment on the compatibility of the above lagrangian formalisms with the high-energy constraints for concrete VVP correlator.Comment: 34 pages, 3 figure

Role of the Acidic Hirudin-like COOH-Terminal Amino Acid Region of Factor Va Heavy Chain in the Enhanced Function of Prothrombinase†

Prothrombinase activates prothrombin through initial cleavage at Arg(320) followed by cleavage at Arg(271). This pathway is characterized by the generation of an enzymatically active, transient intermediate, meizothrombin, that has increased chromogenic substrate activity but poor clotting activity. The heavy chain of factor Va contains an acidic region at the COOH terminus (residues 680-709). We have shown that a pentapeptide from this region (DYDYQ) inhibits prothrombin activation by prothrombinase by inhibiting meizothrombin generation. To ascertain the function of these regions, we have created a mutant recombinant factor V molecule that is missing the last 30 amino acids from the heavy chain (factor V(Delta680-709)) and a mutant molecule with the (695)DYDY (698) --\u3e AAAA substitutions (factor V(4A)). The clotting activities of both recombinant mutant factor Va molecules were impaired compared to the clotting activity of wild-type factor Va (factor Va (Wt)). Using an assay employing purified reagents, we found that prothrombinase assembled with factor Va(Delta680-709) displayed an approximately 39% increase in k cat, while prothrombinase assembled with factor Va(4A) exhibited an approximately 20% increase in k cat for the activation of prothrombin as compared to prothrombinase assembled with factor Va(Wt). Gel electrophoresis analyzing prothrombin activation by prothrombinase assembled with the mutant molecules revealed a delay in prothrombin activation with persistence of meizothrombin. Our data demonstrate that the COOH-terminal region of factor Va heavy chain is indeed crucial for coordinated prothrombin activation by prothrombinase because it regulates meizothrombin cleavage at Arg(271) and suggest that this portion of factor Va is partially responsible for the enhanced procoagulant function of prothrombinase

Single-nucleotide polymorphism associations with preterm delivery: a case-control replication study and meta-analysis

BackgroundThe aim of this study was to replicate single-nucleotide polymorphism (SNP) associations with preterm birth (PTB; birth at MethodsSpontaneous PTB cases and controls were selected from an existing cohort. Candidate SNPs were taken from an existing genotype panel. A systematic review was conducted for each SNP in the panel to determine suitability as a PTB candidate. Those with significant associations previously reported in Caucasians were selected for replication. Candidate SNPs were already genotyped in cases and controls and clinical data were accessed from state perinatal and cerebral palsy databases. Association analysis was conducted between each SNP and PTB, and meta-analysis was conducted if there were ≥ 3 studies in the literature. Maternal and fetal SNPs were considered as separate candidates.ResultsA cohort of 170 cases and 583 controls was formed. Eight SNPs from the original panel of genotyped SNPs were selected as PTB candidates and for replication on the basis of systematic literature review results. In our cohort, fetal factor V Leiden (FVL) was significantly associated with PTB (odds ratio (OR): 2.6, 95% confidence interval (CI): 1.31-5.17), and meta-analysis confirmed this association (OR: 2.71, 95% CI: 1.15-6.4).ConclusionReplication and meta-analysis support an increased risk of PTB in Caucasians with the fetal FVL mutation.Michael E. O’Callaghan, Alastair H. MacLennan, Gai L. McMichael, Eric A. Haan and Gustaaf A. Dekke

Amino Acids Glu323, Tyr324, Glu330, and Val331 of Factor VA Heavy Chain Are Essential for Expression of Cofactor Activity

We have recently demonstrated that amino acid region 323-331 of factor Va heavy chain (9 amino acids, AP4\u27) contains a binding site for factor Xa (Kalafatis, M., and Beck, D. O. (2002) Biochemistry 41, 12715-12728). To ascertain which amino acids within this region are important for the effector and receptor properties of the cofactor with respect to factor Xa, we have synthesized three overlapping peptides (5 amino acids each) spanning the amino acid region 323-331 and tested them for their effect on prothrombinase complex assembly and function. Peptide containing amino acids 323EYFIA327 alone was found to increase the catalytic efficiency of factor Xa but had no effect on the fluorescent anisotropy of active site-labeled factor Xa (human factor Xa labeled in the active site with Oregon Green 488; [OG488]-EGR-hXa). In contrast, peptide containing the sequence 327AAEEV331 was found to interact with [OG488]-EGR-hXa with half-maximal saturation reached at approximately 150 microm, but it was unable to produce a cofactor effect on factor Xa. Peptide 325FIAAE329 inhibited prothrombinase activity and was able to partially decrease the fluorescent anisotropy of [OG488]-EGR-hXa but could not increase the catalytic efficiency of factor Xa with respect to prothrombin. A control peptide with the sequence FFFIA did not increase the catalytic efficiency of factor Xa, whereas a peptide with the sequence AAEMI was impaired in its capability to interact with [OG488]-EGR-hXa. Two mutant recombinant factor Va molecules (Glu323 --\u3e Phe/Tyr324 --\u3e Phe, factor VaFF; Glu330 --\u3e Met/Val331 --\u3e Ile, factor VaMI) showed impaired cofactor activity when used at limiting cofactor concentration, whereas the quadruple mutant (Glu323 --\u3e Phe/Tyr324 --\u3e Phe and Glu330 --\u3e Met/Val331 --\u3e Ile, factor VaFF/MI) had no cofactor activity under similar experimental conditions. Our data demonstrate that amino acid residues Glu323, Tyr324, Glu330, and Val331 of factor Va heavy chain are critical for expression of factor Va cofactor activity

Contribution of Amino Acid Region 334−335 from Factor Va Heavy Chain to the Catalytic Efficiency of Prothrombinase†

ABSTRACT: We have demonstrated that amino acids E323, Y324, E330, and V331 from the factor Va heavy chain are required for the interaction of the cofactor with factor Xa and optimum rates of prothrombin cleavage. We have also shown that amino acid region 332-336 contains residues that are important for cofactor function. Using overlapping peptides, we identified amino acids D334 and Y335 as contributors to cofactor activity. We constructed recombinant factor V molecules with the mutations D334 f K and Y335 f F (factor VKF) and D334 f A and Y335 f A (factor VAA). Kinetic studies showed that while factor VaKF and factor VaAA had a KD for factor Xa similar to the KD observed for wild-type factor Va (factor VaWT), the clotting activities of the mutant molecules were impaired and the kcat of prothrombinase assembled with factor VaKF and factor VaAA was reduced. The second-order rate constant of prothrombinase assembled with factor VaKF or factor VaAA for prothrombin activation was ∼10-fold lower than the second-order rate constant for the same reaction catalyzed by prothrombinase assembled with factor VaWT. We also created quadruple mutants combining mutations in the amino acid region 334–335 with mutations at the previously identified amino acids that are important for factor Xa binding (i.e., E323Y324 and E330V331). Prothrombinase assembled with the quadruple mutant molecules displayed a second-order rate constant up to 400-fold lower than the values obtained with prothrombinase assembled with factor VaWT. The dat

A Control Switch for Prothrombinase: Characterization of a Hirudin-Like Pentapeptide From the COOH Terminus of Factor VA Heavy Chain That Regulates the Rate and Pathway for Prothrombin Activation

Membrane-bound factor Xa alone catalyzes prothrombin activation following initial cleavage at Arg(271) and prethrombin 2 formation (pre2 pathway). Factor Va directs prothrombin activation by factor Xa through the meizothrombin pathway, characterized by initial cleavage at Arg(320) (meizo pathway). We have shown previously that a pentapeptide encompassing amino acid sequence 695-699 from the COOH terminus of the heavy chain of factor Va (Asp-Tyr-Asp-Tyr-Gln, DYDYQ) inhibits prothrombin activation by prothrombinase in a competitive manner with respect to substrate. To understand the mechanism of inhibition of thrombin formation by DYDYQ, we have studied prothrombin activation by gel electrophoresis. Titration of plasma-derived prothrombin activation by prothrombinase, with increasing concentrations of peptide, resulted in complete inhibition of the meizo pathway. However, thrombin formation still occurred through the pre2 pathway. These data demonstrate that the peptide preferentially inhibits initial cleavage of prothrombin by prothrombinase at Arg(320). These findings were corroborated by studying the activation of recombinant mutant prothrombin molecules rMZ-II (R155A/R284A/R271A) and rP2-II (R155A/R284A/R320A) which can be only cleaved at Arg(320) and Arg(271), respectively. Cleavage of rMZ-II by prothrombinase was completely inhibited by low concentrations of DYDYQ, whereas high concentrations of pentapeptide were required to inhibit cleavage of rP2-II. The pentapeptide also interfered with prothrombin cleavage by membrane-bound factor Xa alone in the absence of factor Va increasing the rate for cleavage at Arg(271) of plasma-derived prothrombin or rP2-II. Our data demonstrate that pentapeptide DYDYQ has opposing effects on membrane-bound factor Xa for prothrombin cleavage, depending on the incorporation of factor Va in prothrombinase

Contribution of Amino Acid Region 659−663 of Factor Va Heavy Chain to the Activity of Factor Xa within Prothrombinase†,‡

ABSTRACT: Factor Va, the cofactor of prothrombinase, is composed of heavy and light chains associated noncovalently in the presence of divalent metal ions. The COOH-terminal region of the heavy chain contains acidic amino acid clusters that are important for cofactor activity. In this work, we have investigated the role of amino acid region 659-663, which contains five consecutive acidic amino acid residues, by site-directed mutagenesis. We have generated factor V molecules in which all residues were mutated to either lysine (factor V 5K) or alanine (factor V 5A). We have also constructed a mutant molecule with this region deleted (factor V Δ659-663). The recombinant molecules along with wild-type factor V (factor V WT) were transiently expressed in mammalian cells, purified, and assessed for cofactor activity. Two-stage clotting assays revealed that the mutant molecules had reduced clotting activities compared to that of factor Va WT. Kinetic analyses of prothrombinase assembled with the mutant molecules demonstrated diminished k cat values, while the affinity of all mutant molecules for factor Xa was similar to that for factor Va WT. Gel electrophoresis analyses of plasma-derived and recombinant mutant prothrombin activation demonstrated delayed cleavage of prothrombin at both Arg 320 and Arg 271 by prothrombinase assembled with the mutant molecules, resulting i