Ceruloplasmin and myeloperoxidase in complex affect the enzymatic properties of each other

Ceruloplasmin (CP), the multicopper oxidase of plasma, interacts with myeloperoxidase (MPO), an enzyme of leukocytes, and inhibits its peroxidase and chlorinating activity. Studies on the enzymatic properties shows that CP behaves as a competitive inhibitor impeding the binding of aromatic substrates to the active centre of MPO. The contact between CP and MPO probably entails conformational changes close to the p-phenylenediamine binding site in CP, which explains the observed activation by MPO of the substrate's oxidation. CP subjected to partial proteolysis was virtually unable to inhibit activity of MPO. The possible protein–protein interface is comprised of the area near active site of MPO and the loop linking domains 5 and 6 in CP. One of the outcomes of this study is the finding of a new link between antioxidant properties of CP and its susceptibility to proteolysis.     

Study of Interaction of Ceruloplasmin with Serprocidins

This paper describes formation of complexes of ceruloplasmin (CP) with such proteins of the serprocidin family as azurocidin (CAP37), neutrophilic elastase (NE), cathepsin G (CG), and proteinase 3 (PR3). We present evidence that serprocidins form complexes with CP at a molar ratio 1: 1. Phenylmethylsulfonyl fluoride, a serine protease inhibitor, did not prevent the interaction of serprocidins with CP in the course of SDS-free disc electrophoresis. CP affected the activities of NE, CG, and PR3 as a competitive inhibitor with K _i ≈ 1 μM. Inhibitory effect of CP depended on ionic strength of the solution and was negligible at NaCl concentrations above 300 mM. In the mode of competitive inhibitors serprocidins suppressed oxidase activity of CP towards p-phenylenediamine. CAP37 displayed the strongest inhibitory effect (K _i ≈20 nM). Upon adding various serprocidins to human, rat, rabbit, dolphin, dog, horse, and mouse plasma only CAP37 would form a complex with CP. Synthetic peptide RKARPRQFPRRR (5–13, 61–63 CAP37) displaced CAP37 from its complex with CP. Adding CAP37 to the triple complex formed by CP, lactoferrin, and myeloperoxidase resulted in displacement of the latter from the complex. The dissociation constant of CAP37 with immobilized CP was 13 nM. Therefore, among serprocidins CAP37 can be regarded as the specific partner of CP.     

Plant fiber intrusive growth characterized by NMR method

Intrusively growing plant cells insert themselves between surrounding cells, thus increasing the number of membranes on the tissue cross-section. This parameter can be assessed by spin echo NMR method with a magnetic field pulse gradient. Diffusion echo decay was measured for stem regions of long-fiber flax (Linum usitatissimum L.) differing in the stages of primary fiber development, which elongate thousand-fold during intrusive growth. Additionally, the number of fibers on stem cross-sections was counted under microscope. An increase in the slow component of the echo diffusion decay was correlated with an increase in the number of fibers on the stem cross-section in the zone of intrusive growth, while other stem-structure characteristics remained unchanged. Thus, NMR method can be used for characterization of intrusive fiber growth in situ.     

Identification and properties of complexes formed by myeloperoxidase with lipoproteins and ceruloplasmin

The first evidence of multi-component complexes formed by myeloperoxidase (MPO), ceruloplasmin (CP), and very low/low density lipoproteins (VLDL/LDL) obtained by electrophoresis, gel filtration, and photon-correlation spectroscopy (PCS) is presented in this paper. Complexes were observed when isolated MPO, CP, and VLDL/LDL were mixed and/or when MPO was added to the blood plasma. Complex LDL–MPO–CP was detected in 44 of 100 plasma samples taken from patients with atherosclerosis, and 33 of 44 samples also contained the VLDL–MPO–CP complex. MPO concentration in these patients’ plasma exceeded 800 ng/ml. Interaction of MPO with high density lipoproteins (HDL) was not revealed, as well as binding of CP to lipoproteins in the absence of MPO. Adding antibodies against apoB-100 to VLDL–MPO–CP and LDL–MPO–CP complexes results in release of lipoproteins. Using PCS the diameters of complexes under study were evaluated. By comparing concentrations of the components in complexes formed by MPO, CP, and lipoproteins their stoichiometry was assessed as 2VLDL:1MPO:2CP and 1LDL:1MPO:2CP. Lipoproteins affected the inhibition of MPO peroxidase activity by CP. The affinity of lipoproteins to MPO–CP complex was assessed using apparent dissociation constants determined as ～0.3 nM for VLDL and ～0.14 nM for LDL.     

Reactogenicity and immunological effectiveness study of a complex paratyphoid B antigen]

Reactogenic property and immunological efficacy of the paratyphoid preparation containing a complex of O-, K- and H-antigens obtained by single-stage antigens extraction were studied in a limited group of volunteers (22 persons). The antigen gave no untoward reactions and proved to be safe when given orally in doses of 25 to 150 mg. Paratyphoid B antigen was characterized by a marked immunization activity and stimulated formation of specific paratyphoid O-, K- and H-agglutinins and antibodies of the IgA,- IgG,- and IgM-classes.     

The role of cellular and humoral factors of body resistance in the development of experimental influenza-staphylococcal infections in mice

The clinico-experimental studies of mixed influenza-staphylococcal infection constantly point to the development of the aggravation of the infectious process due to the synergic action of the bacterial and viral infective agents. But, as shown by the authors of the present work, in those cases when the experimental infection with the virus was preceded by staphylococcal infection by 72 hours no synergism was observed. In cases of infection with adaptogenic virus the mortality rate of mice resulting from meningococcal infection was twice as low. The possible explanation of this fact is discussed.     

Effect of lactoferrin on oxidative features of ceruloplasmin

In our previous report we first described a complex between lactoferrin (Lf) and ceruloplasmin (Cp) with K _d ~ 1.8 μM. The presence of this complex in colostrum that never contains more than 0.3 μM Cp questions the reliability of K _d value. We carefully studied Lf binding to Cp and investigated the enzymatic activity of the latter in the presence of Lf, which allowed obtaining a new value for K _d of Cp–Lf complex. Lf interacting with Cp changes its oxidizing activity with various substrates, such as Fe²⁺, o-dianisidine (o-DA), p-phenylenediamine (p-PD) and dihydroxyphenylalanine (DOPA). The presence of at least two binding sites for Lf in Cp molecule is deduced from comparison of substrates’ oxidation kinetics with and without Lf. When Lf binds to the first site affinity of Cp to Fe²⁺ and to o-DA increases, but it decreases towards DOPA and remains unchanged towards p-PD. Oxidation rate of Fe²⁺ grows, while that of o-DA, p-PD and DOPA goes down. Subsequent Lf binding to the second center has no effect on iron oxidation, hampers DOPA and o-DA oxidation, and reduces affinity towards p-PD. Scatchard plot for Lf sorbing to Cp-Sepharose allowed estimating K _d for Lf binding to high-affinity (~13.4 nM) and low-affinity (~211 nM) sites. The observed effect of Lf on ferroxidase activity of Cp is likely to have physiological implications.     

Pathogen‐induced conditioning of the primary xylem vessels – a prerequisite for the formation of bacterial emboli by Pectobacterium atrosepticum

Representatives of Pectobacterium genus are some of the most harmful phytopathogens in the world. In the present study, we have elucidated novel aspects of plant–Pectobacterium atrosepticum interactions. This bacterium was recently demonstrated to form specific ‘multicellular’ structures – bacterial emboli in the xylem vessels of infected plants. In our work, we showed that the process of formation of these structures includes the pathogen‐induced reactions of the plant. The colonisation of the plant by P. atrosepticum is coupled with the release of a pectic polysaccharide, rhamnogalacturonan I, into the vessel lumen from the plant cell wall. This polysaccharide gives rise to a gel that serves as a matrix for bacterial emboli. P. atrosepticum‐caused infection involves an increase of reactive oxygen species (ROS) levels in the vessels, creating the conditions for the scission of polysaccharides and modification of plant cell wall composition. Both the release of rhamnogalacturonan I and the increase in ROS precede colonisation of the vessels by bacteria and occur only in the primary xylem vessels, the same as the subsequent formation of bacterial emboli. Since the appearance of rhamnogalacturonan I and increase in ROS levels do not hamper the bacterial cells and form a basis for the assembly of bacterial emboli, these reactions may be regarded as part of the susceptible response of the plant. Bacterial emboli thus represent the products of host–pathogen integration, since the formation of these structures requires the action of both partners.     

A model for the study of the mechanism of a low pH-induced interaction of the virus fusion proteins and cell membranes

A model is proposed for the study of molecular mechanisms of a low pH-induced interaction of fusion proteins of enveloped viruses and cell membranes. The model consists of large monolamellar liposomes containing ionophore nigericin in their membranes and ectodomains of fusion protein in their inner space. The process of interaction of the protein with the lipid bilayer is triggered by acidification of the liposomal constituents to the pH of fusion with the help of nigericin by adding citric acid to the outer medium. To visualize the protein structural reorganization, the tritium planigraphy was used.Comparison of the values of specific labelling of the proteins and distribution of radioactivity in individual amino acids in control (at neutral pH) and experimental liposome samples (at the pH of fusion) permits to realise the character of protein-membrane interaction. We have obtained the first results in the study of interaction of the bromelain-released soluble ectodomain of the HA^XX molecule (BHA)—with the lipid membrane. The observed increase in the protein specific activity and selective increase in the specific activity of hydrophobic amino acids Ile, Phe and Tyr in experimental liposome samples as compared with the controls did not contradict to the conventional concept, that a hydrophobic N-terminus of HA2 subunit of hemagglutinin is responsible for its interaction with lipid membranes.