Examination of protein sequence homologies: III. Ribosomal protein YS25 fromSaccharomyces cerevisiae and its counterparts fromSchizosaccharomyces pombe,rat liver,andEscherichia coli

Summary The sequences of the ribosomal proteins YS25, SP-S28, RL-S21, and Ec-S6, fromSaccharomyces cerevisiae, Schizosaccharomyces pombe, rat liver, andEscherichia coli, respectively, have been examined using a computer program that searches for homologous tertiary structures. Matrices of comparisons among the eukaryotic sequences show that they match each other sequentially without any internal gaps. The average values of the correlation coefficients obtained from the comparison matrices are higher for the first halves of the sequences than for the latter halves. This result suggests that the first halves of the sequences may represent a more important domain than the latter halves. The comparison matrices between the eukaryotic and bacterial sequences of ribosomal proteins, however, do not show sequentially arranged homology, though there are six well-matching segments arranged in different orders in the two types of sequences. This implies that the eukaryotic sequences of the ribosomal protein were reconstituted by two internal transpositions and six deletions of 4–12 residues each from the ancestral sequence during the divergence between bacterial and eukaryotic genes. These findings may give insight into structural and quantitative studies of evolutionary divergence between eukaryotes and prokaryotes.     

Characteristics of a Urate-Degrading Diamine Oxidase-Peroxidase Enzyme System in Soybean Radicles

The cadaverine content of soybean radicles showed a maximumpeak 3–4 days after planting. The variation coincidedwith radicle uricase activity during seed germination. The uricase activity could not be fractionate when the bufferpH for the extraction was at 6.0. The addition of 1 M KCl orNaCl to the buffer allowed the extraction of the uricase activity,but an addition of 1 M MgCl₂ or BaCl₂ inhibited this enzyme'sactivity. The urate-degrading enzyme system was purified 248-fold permilligram of protein from soybean radicles. The respective K_mvalues of the diamine oxidase activity for cadaverine and ofthe urate-degrading activity for hydrogen peroxide and uratewere 1.25, 2.93 and 50.3 µM. Analysis by gel electrophoresisof the partially purified enzyme fraction revealed that theurate-degrading enzyme system consisted of a peroxidase thatdegrades urate with hydrogen peroxide and a diamine oxidasethat releases hydrogen peroxide. These data are evidence that a urate-degrading diamine oxidaseand peroxidase system exists in soybean radicles and that thereaction rate of urate-degradation is controlled by the concentrationof cadaverine. (Received November 28, 1984; Accepted April 8, 1985)     

Fibrinogen distribution on surfaces and in organelles of ADP stimulated human blood platelets

The fibrinogen distribution in platelet organelles after ADP-stimulation was investigated with anti-human fibrinogen using protein A-gold applied to serial sections. Fibrinogen was detected in the so-called alpha-granules of platelets and also in granule protrusions which were observed after ADP-stimulation. The ends of these protrusions were formed as coated membranes and the tips were often in apposition to the surface connected membranes or the plasmalemma. At such places fusion events and hence signs of an exocytosis could be demonstrated by means of cryofixation and cryosubstitution. Examination of serial sections revealed fibrinogen on all these granule profiles. Surface connected membranes, free surfaces and the characteristic structure of the contact zones of aggregated platelets were also labelled by gold particles but less than anticipated. On the platelet surfaces and surface connected membranes fibrinogen was rarely demonstrable with ferritin-labelled anti-human fibrinogen on washed or thrombin-stimulated, almost fibrinogen free platelets. After addition of human fibrinogen to the thrombin stimulated and disaggregated platelets a part of the platelets aggregated spontaneously and formed characteristic contact zones. Anti-human fibrinogen was observed on the free surfaces, in filamentous bridges between the contact spaces and in a tubular surface connected membrane system with involvement of coated membranes at the central ends of these structures. The results indicate the following: all alpha-granules contain fibrinogen; after ADP-stimulation secretion takes place with involvement of coated membranes; during aggregation fibrinogen binds to platelet surfaces and forms contact spaces; fibrinogen is taken up by the surface connected system with involvement of coated membranes.     

Serological survey of T-lymphocyte differentiation antigens in wild mice

Allelic distributions of Thy-1, Ly-l, and Ly-2 antigens in wild mice are characteristic of each Mus musculus subspecies. Eastern mice (M.m.molossinus, M.mmusculus, M.m.castaneus, M.m.bactrianus) express the Thy-1.1 antigen, whereas Western mice (M.m. domesticus, M.m.brevirostris) express the Thy-1.2. All mice from wild populations examined in this survey express the Ly-1.2. The Ly-2.1 is distributed in Eastern mice and some Western mice, and the Ly-2.2 is found in the remaining Western mice. Allelic distributions of these antigens were also examined in two other species, Mus spretus and Mus spicilegus. Allelic constitutions of Thy-1 and Ly-1 in these species are similar to those of Eastern mice. Some M.spicilegus, however, express the Ly-1.1 antigen. This antigenic type is not found in M.musculus. Some Eastern mice related to M.m.castaneus react weakly to Ly-1.2-specific and Ly-2.1-specific monoclonal antibodies in both the complement-mediated cytotoxicity test and the absorption test. These results suggest that M.m.castaneus has unique alleles in the Ly-1 and Ly-2 loci.     

Blood Types in Relation to Depressions and Schizophrenia: A Preliminary Report

The distribution of the ABO, A₁, D, E, C, and Kell blood types in relation to selected psychiatric diagnoses was studied in over 700 consecutive admissions to a provincial mental hospital. Predicted relationships based upon recent work in other laboratories were supported by consistent trends approaching statistical significance. In addition, blood group O was found to be significantly associated with involutional melancholia.     

Characterization of human thyrotropin receptor-related peptide-like immunoreactivity in peripheral blood of Graves' disease.

An antiserum raised against an alignment of amino acid-(32-56), termed TSHRP-1, in the extracellular domain of human thyrotropin (TSH) receptor was used to identify the TSH receptor-like substance in plasma of Graves' disease. The dilution curve of plasma TSHRP-1-like immunoreactivity was observed in a manner parallel to the standard synthetic peptide curve in radioimmunoassay, and its molecular weight estimated approximately 60 kDa. The amounts of TSHRP-1-like immunoreactivity were significantly higher in Graves' plasma than those in plasma of normal and hypothyroid patients due to Hashimoto's thyroiditis. The present results indicate that human peripheral blood possesses a soluble form of the extracellular domain of TSH receptor which may contribute to the pathophysiology of Graves' disease.     

Organization and expression inPseudomonas putida of the gene cluster involved in cephalosporin biosynthesis fromLysobacter lactamgenus YK90

TheLysobacter lactamgenus YK90pcbAB gene encoding -(l--aminoadipyl)-l-cysteinyl-d-valine (ACV) synthetase is located immediately upstream of thepcbC gene in the same orientation in the gene cluster involved in cephalosporin biosynthesis. ThepcbAB gene encodes a large polypeptide composed of 3722 amino acid residues with a molecular mass of 411 593 Da. The predicted amino acid sequence has a high degree of similarity with those of known ACV synthetases from fungi and actinomycetes. Within thepcbAB amino acid sequence, three conserved and repeated domains of about 600 amino acids were identified. The domains also share a high degree of similarity with non-ribosomal peptide synthetases such as gramicidin synthatase 2 ofBacillus brevis. ThepcbAB gene was expressed under the control of thelac promoter inPseudomonas putida. Expression of the gene cluster involved in cephalosporin biosynthesis inP. putida led to the accumulation of -lactam antibiotics. Deletion analysis of an open-reading frame located between thecefE andcefD genes from the gene cluster revealed that it encoded deacetylcephalosporin C synthetase (cefF). From the results presented here and those of previous studies, the genes involved in cephalosporin biosynthesis inL. lactamgenus appear to be clustered in the orderpcb AB-pcbC- cefE-cefF-cefD-bla in the same orientation within a 17-kb region of DNA.     

Evaluation of the Sensitivity and Specificity of Polymerase Chain Reaction Test Kit,AMPLICOR Chlamydia trachomatis

The sensitivity and specificity of the polymerase chain reaction (PCR) test kit, AMPLICOR Chlamydia trachomatis, were examined by the use of purified elementary bodies (EBs), cells having inclusions containing reticulate bodies alone and 20 clinical isolates. The numbers of EB and inclusion of C. trachomatis at the detection limit were determined to be approximately 2 to 4 EBs and one inclusion per assay, respectively. No reaction occurred for C. psittaci and C. pneumoniae. All clinical isolates were positively reacted in the PCR assay.