The polypeptide structure and assembly of Ly-2/3 heterodimers

Mild reduction of mature, thymic Ly-2/3 heterodimers of M _r 67 000 resulted in dissociation into three individual polypeptide chains, , , and , of respective M _r values 38000, 35000, and 30000. The and chains were both immunoprecipitated by a monoclonal antibody directed to the Ly-2.1 epitope whereas the Ly-3.1 antibody bound only the chain. The possibility that the and chains of each heterodimer established their interchain links within a labile precursor protein in which a and segments were fused was considered but discounted by the finding that in mice heterozygous for both Ly-2 and Ly-3 loci, the Ly-2 product of one chromosome was not exclusively joined to Ly-3 structures coded by the same chromosome. By utilizing ionic detergents which selectively alter the charge of intrinsic membrane proteins, both Ly-2 and Ly-3 polypeptides were shown to have membrane insertion sites. It is suggested that as a consequence of their likely synthesis on membrane-bound polysomes, newly synthesized Ly-2 and Ly-3 structures accumulate within the same subcellular compartment — the membranes of the rough endoplasmic reticulum. Their elevated concentration within this space may facilitate a low affinity binding interaction between Ly-2 and Ly-3 which is later stabilized by interchain disulfide bond formation.Abbreviations used in this paper BSA bovine serum albumin - DOC sodium deoxycholate - DTT dithiothreitol - HA hemagglutinin - HTAB hexadecyltrimethylammonium bromide - RER rough endoplasmic reticulum - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - TX100 Triton X-100     

Correlated changes in steady-state levels of Balbiani ring mRNAs and secretory polypeptides in salivary glands of Chironomus tentans

Balbiani rings (BRs) on polytenized chromosomes in Chironomid salivary glands contain members of a homologous multigene family that encodes a family (the sp-I family) of high M _r secretory polypeptides. Each of these BR genes is comprised largely of tandemly duplicated core repeat sequences consisting of related constant (C) regions and intergenically divergent subrepeat (SR) regions. A set of oligodeoxyribonucleotide probes were synthesized that correspond to the transcribed strand of the SR region of BR1, BR2, BR2/, and BR6 core repeats. Under a defined set of conditions, it was possible to show that each oligonucleotide probe hybridized exclusively to its cognate repeat type without hybridization to other repeat types in cloned DNA templates. These BR probes were then used in dotblot hybridization experiments to simultaneously follow alterations in the steady-state level of BR mRNAs in response to prolonged exposure of larvae to galactose. The results indicated that the relative amounts of these four BR mRNAs may change in a noncoordinate manner. These BR probes were also used in experiments to compare simultaneously the salivary gland content of sp-I components and specific BR mRNAs in larvae that exhibited naturally occurring or induced alterations in BR gene expression. A correlation was found which suggested that sp-Ia is encoded in a gene comprised of BR1 repeats, sp-Ib is encoded by BR2 repeats, sp-Ic is encoded by BR6 repeats and sp-Id is encoded by BR2 repeats.     

A new member of the Balbiani ring multigene family in the dipteran Chironomus tentans consists of a single-copy version of a unit repeated in other gene family members

The known Balbiani ring (BR) multigene family members in the dipteran Chironomus tentans encode salivary gland secretory proteins in the size range between 38 and 1,000 kDa. The proteins interact to form protein fibers used by the aquatic larvae to spin feeding and protective larval tubes or pupation tubes. Here, we describe a new BR multigene family member, the spl7 gene, which codes for an 89-amino-acid-long protein with a relative mobility of 17k. The gene has a high content of charged amino acid residues and consists of two structurally different halves. Five regularly spaced cysteine codons are present in the 5 half while the 3 half contains five proline codons. These two different halves exhibit similarities to the C and SR regions, respectively, which form the tandemly repeated units in the about 40-kb-long BR genes and which also, in different versions, are the building blocks of all genes in the BR multigene family.In this multigene family, encoding interacting structural proteins, the long BR genes with their 125–150 tandemly arranged repeat units as well as the short sp17 gene with its single-copy version of such a repeat unit, have therefore evolved from a common ancestor.Correspondence to: L. Wieslander     

Is There a Field-Theoretic Explanation For Precursor Biopolymers?

A Hu-Barkana-Gruzinov cold dark matter scalarfield may enter a weak isospin invariant derivative interactionthat causes the flow of right-handed electrons to align parallelto (). Hence, in the outer regions of galaxies where () islarge, as in galactic halos, the derivative interaction mayinduce a chirality-imbued quantum chemistry. Such a chirality-imbued chemistry would in turn be conducive to the formation ofabundant precursor biopolymers on interstellar dust grains,comets and meteors in galactic halo regions, with subsequentdelivery to planets in the inner galactic regions where and() are concomitantly near zero and left-right symmetricterrestrial quantum chemistry prevails.     

Ultrastructure of four types of striated muscle fibers in the atlantic hagfish (Myxine glutinosa,L.)

Summary Four types of striated muscle fibers with distinctive ultrastructure were defined in the Atlantic hagfish (Myxine glutinosa, L.): white, intermediate, and red fibers of m. parietalis, and red fibers of m. craniovelaris.White fibers are thick, contain very few mitochondria and fat vacuoles, and possess distinct and separate myofibrils with thin Z-disks and distinct M-lines. Intermediate fibers are thinner, possess largely similar myofibrils that often are even better separated due to a higher content of fat vacuoles and especially mitochondria and glycogen granules. Red fibers of m. parietalis contain large amounts of mitochondria, fat vacuoles, and glycogen granules. Their myofibrils possess M-lines, and although branching more, the myofibrils of red fibers conform with a Fibrillenstruktur pattern like those of white and intermediate fibers. Red fibers of m. craniovelaris are very thin, possess many smaller fat vacuoles, and large amounts of mitochondria and glycogen granules. The myofibrils are significantly thinner than in m. parietalis fibers, run as quite independent well separated units, possess thicker Z-disks, and lack M-lines. Large amounts of myosatellite cells are associated with these red fibers.Triads are located near A/I-junctions in all four fiber types and occur irregularly, the density of triads being different in the various fiber types.We are indebted to Dr. Finn Walvig, Biological Station, University of Oslo, Drøbak, for supply of hagfishes, and we also wish to thank Dr. Jan K. S. Jansen, Institute of Physiology, University of Oslo, for valuable suggestions during this study.     

High crossability of wild barley (Hordeum spontaneum C. Koch) with bread wheat and the differential elimination of barley chromosomes in the hybrids

Four bread wheat (Triticum aestivum L.) cultivars, Aobakomugi, Chinese Spring, Norin 61 and Shinchunaga, were pollinated with five barley lines/cultivars consisting of three cultivated barley (Hordeum vulgare L.) lines, Betzes, Kinai 5 and OHL089, and two wild barley (Hordeum spontaneum C. Koch) lines, OUH602 and OUH324. Crossability, expressed as the percentage of embryo formation, varied from 0 to 55.4% among the cross combinations. The two wild barley lines generally had a higher crossability than the previously reported best pollinator, Betzes, and some Japanese wheat cultivars were better as the female parent than Chinese Spring. Ninety four hybrid plants were obtained from 250 embryos cultured, and their somatic chromosome numbers ranged from 21 to 36. Eighteen plants were mosaic in chromosome number. Twenty one-chromosome plants appeared most frequently (45.7%) followed by 28-chromosome plants (14.9%). C-banding analysis revealed that elimination of barley chromosomes was mainly responsible for the occurrence of aneuploid plants. In hypoploids derived from Betzes-crosses, chromosome 5 was preferentially eliminated as previously reported, while in hypoploids derived from OUH602-crosses, chromosome 4 was preferentially eliminated. The wild barley line OUH602 may be a useful parent for producing a new wheat-barley addition set because of its high crossability with wheat and a different pattern of chromosome elimination.     

γ-Glutamylamine cyclotransferase

Summary -Glutamylamine cyclotransferase, an enzyme found in a number of animal tissues and cells, catalyzes the conversion of -(L--glutamyl)-L-lysine to free lysine and 5-oxo-L-proline as well as the release of free amines and the formation of 5-oxo-L-proline from a variety of other L--glutamylamines. Among its substrates are both the mono- and di--glutamyl derivatives of putrescine, spermidine and spermine, and a derivative of -(L--glutamyl)-L-lysine in which both the -amino group and the carboxyl group of the lysine moiety are blocked. The enzyme does not act on most -glutamyl--amino acids, nor is it active toward the -lysyl derivatives of L-aspartic acid or D-glutamic acid. Derivatives of -(L--glutamyl)-L-lysine in which the -amino or the -carboxyl function of the glutamyl moiety is blocked also do not serve as substrates. The specificity of -glutamylamine cyclotransferase is in accordance with the proposal that it functions biologically in the latter stages of the catabolism of products of the action of transglutaminases. Some suggestions as to the manner in which -glutamylamine cyclotransferase serves this function are made based on present knowledge of protein degradation.     

Structural implications of primary sequences from a family of Balbiani ring-encoded proteins inChironomus

Summary DNA sequencing has revealed an internal, tandemly repetitive structure in the family of giant polypeptides encoded by three types of Balbiani ring (BR) genes, in three different species ofChironomus. Each major BR repeat can be subdivided into two halves: a region consisting of short subrepeats and a more constant region that lacks obvious subrepeats. Comparative predictions of secondary structure indicate that an -helical segment is consistently present in the amino-terminal half of the constant region in all known BR proteins. Comparative predictions, coupled with consideration of the known phosphorylation of serine and threonine residues in BR proteins, suggest that the -helical structure may also extend into the carboxy-terminal half of the constant region, possibly interrupted by -turn(s). However, it is also possible that the structure is variable, and that a -strand is present in that half in some cases. All of the constant regions conserve one methionine and one phenylalanine residue, as well as all four cysteines; these residues presumably play roles in the packing or cross-linking of aligned constant regions. The structure of the subrepeat region is not clear, but the prevalence of a tripeptide pattern (basic-proline-acidic) suggests some type of structural regularity, possibly an extended helix. The possible significance of these conserved molecular features is discussed in the context of how they may serve the elasticity, insolubility, and hydrophilicity of the fibrils and threads formed by the BR polypeptides.     

Liquid chromatography “on-flow” 1H nuclear magnetic resonance on native glycosphingolipid mixtures together with gas chromatography/mass spectrometry on the released oligosaccharides for screening and characterisation of carbohydrate-based antigens from pig lungs

Glycosphingolipids were prepared from pig lung and pooled into two fractions with (i) 3 sugar residues, and (ii) 3 sugar residues. Oligosaccharides were prepared and used for gas chromatography, gas chromatography-mass spectrometry and matrix-assisted laser desorption/ionization mass spectrometry. The glycolipid fractions i and ii were further characterised and purified using a novel method based on high performance liquid chromatography on-flow proton nuclear magnetic resonance. The LC on-flow NMR technique showed good chromatographic separation and gave NMR spectral information which could be used as guidance for pooling of the separated mixture glycolipids. Conventional ¹H NMR, thin layer immunostaining, gas chromatography, gas chromatography/mass spectrometry and matrix-assisted laser desorption/ionization mass spectrometry were used to characterise the glycolipids and to validate LC-NMR spectral data.     

Modification of length, hydrophobic properties and electric charge of Bacillus subtilis alpha-amylase signal peptide and their different effects on the production of secretory proteins in B. subtilis and Escherichia coli cells

Summary Bacillus subtilis -amylase signal peptide, which consists of 33 amino acids, is functional in Escherichia coli cells.Lysine, glutamic acid, leucine, leucyl-leucine, or leucyl-leucyl-leucine was inserted between positions 28 and 29 of the -amylase signal peptide using site directed mutagenesis. DNAs encoding the wild-type and modified signal peptides were then fused in-frame to DNAs encoding the mature regions of the -lactamase of pBR322 and a thermostable -amylase. The secretion of -lactamase in E. coli cells was more inhibited by the modified signal peptides than that in B. subtilis cells, although the degree of inhibition varied and the inhibitory effect of each signal peptide was found to be similar in the two strains. In contrast, the difference in the inhibitory effect of each modified signal peptide was no longer detected in the case of the production of thermostable -amylase, except for the insertion of glutamic acid. Nearly 50% of thermostable -amylase in the precursor form was accumulated in the intracellular fraction of E. coli cells containing the DNAs for the modified signal peptides. The insertion of glutamic acid inhibited the secretion of the two enzymes in both B. subtilis and E. coli cells.     

Mechanistic possibilities in prebiotic thiophosphate chemistry

Summary Two types of reactivities of thiophosphates have been demonstrated: one being nucleophilic displacement by the P-S moiety of nucleoside phosphorothioates and the other, phosphorylation via P-S cleavage as the driving force. We have designed a system where both displacement on carbon and P-S cleavage are possible. Adenosine derivatives have been synthesized with 5-deoxy-5-chloro and 5-O-tosyl substitutions as leaving groups utilizing the 3-O-phosphorothioate as the biphilic center. The main products of cyclization were 5-O-tosyl and 5-chloroadenosine 2:3-cyclic phosphate. Formation of 3:5-S-phosphorothioate was slow even using an excellent leaving group. This is possibly due to hydrogen bonding between the 2-OH and the neighboring P-O.^– KOH hydrolysis of the cyclic phosphorothioate yielded 2(3) phosphorothioates in a 1:1 ratio. The 2 and 3 isomers were separated and used to study the relative rates of cyclization. The cyclization via P-S cleavage of 2(3)-O-phosphorothioates showed that the 2 isomer was more reactive. This is the first report of superior reactivity of the 3-OH of a ribonucleoside.     

Structural analyses of new tri- and tetrasaccharides produced from disaccharides by transglycosylation of purified Trichoderma viride β-glucosidase

A new -glucosidase was partially purified from Trichoderma viride cellulase. This -glucosidase catalyzed a transglycosylation reaction of cellobiose to give -D-Glc-(16)--D-Glc-(14)-D-Glc (1, yield: 18.8%) and -D-Glc-(16)--D-Glc-(16)--D-Glc-(14)-D-Glc (2, 3.7%), regioselectively. Furthermore, the enzyme regioselectively converted laminaribiose and gentiobiose into -D-Glc-(16)--D-Glc-(13)-D-Glc (3, 15.3%) and -D-Glc-(16)--D-Glc-(16)-D-Glc (4, 20.2%), respectively. The structures (1–4) of the products were determined by ¹H and ¹³C NMR spectroscopies. This high regio- and stereoselectively of the -glucosidase could be applied for oligosaccharide synthesis.     

Conserved and variable repeat structures in the Balbiani ring gene family in Chironomus tentans

Summary The four Balbiani ring (BR) genes, BR1, BR2.1, BR2.2, and BR6 in the midge Chironomus tentans constitute a gene family encoding secretory proteins with molecular weights of approximately 10⁶ daltons. The major part of each gene is known to consist of tandemly organized composite repeat units resulting in a hierarchic repeat arrangement.Here, we present the sequence organization of the 5 part of the BR2.2 and BR6 genes and describe the entire transcribed part of the two genes. As the BR1 and BR2.1 genes were also fully characterized recently, this allows the comparison of all genes in the BR gene family.All four genes share the same exon-intron structure and have evolved by gene duplications starting from a common ancestor, having the same overall organization as the BR genes of today.The genes encode proteins that have an approximately 10,000-amino acid residue extended central domain, flanked by a highly charged, 200-residue amino-terminal domain and a globular 110-residue carboxy-terminal domain. Exons 1–3 and the beginning of exon 4 encode the amino-terminal domain, which throughout contains many regions built from short repeats. These repeats are often degenerate as to repeat unit and sequence and are present in different numbers between the genes. In several instances these repeat structures, however, are conserved at the protein level where they form positively or negatively charged regions.Each BR gene has a 26–38-kb-long exon 4, which consists of an array of 125–150 repeat units and encodes the central domain. The number of repeat units appears to be largely preserved by selection and all repeat units in the array are very efficiently homogenized. Occasionally variant repeats have been introduced, presumably from another BR gene by gene conversion, and spread within the array.Introns 1–3 at the 5 end of the genes have diverged extensively in sequence and length between the genes. In contrast, intron 4 at the 3 end is virtually identical between three of the four genes, suggesting that gene conversion homogenizes the 3 ends of the genes, but not the 5 ends. Offprint requests to: L. Wieslander     

Komparative Aussagemuster in Bezug zu komplementären und enkaptischen Modellen der Morphologie

The unity of organisms can be viewed in terms of the concepts of enkapsis and complementarity. A model (or a type) represents those properties (of elements, structure, and system) which renders cases - the organisms under consideration — comparable. Comparability is established by operations (or metamorphoses) which relate a case to a model. Therefore, the model and the operations must be enumerated together, if a certain morphology is to be established and applied. Two models, which in some way are related, are conjunct, otherwise they are disjunct. If one model is deducible from the other, they are enkaptic conjunct. If the models are essentially different, that is to say that they cannot be transduced into each other, although they condition each other, they are complementary conjunct: although not comparable themselves, only both together describe a case (or a set of cases) completely. Now, the comparability of a case with two models is considered. The two basic patterns of general comparability are homology and analogy. If the two models are complementary conjunct, nine patterns of special comparability can be distinguished. Each is named in accordance with the general meaning of homology and analogy and, as far as possible, with conventional scientific usage. With minor modifications the terminology also applies to more complicated patterns of comparability including distinct or different conjunct models.     

Relationship between transport and metabolism of α-naphthaleneacetic acid, β-naphthaleneacetic acid and α-decalylacetic acid in segments of Coleus

Summary Transportand metabolism of -naphthaleneacetic acid -naphthaleneacetic acid, and -decalylacetic acid, all labelled with ¹⁴C in the carboxyl, group, were studied. Only -naphthaleneacetic acid is transported in a polar way. Most of the radioactivity in the tissue is in a low molecular form, either free or as immobilization products. The immobilization of -naphthaleneacetic acid is similar to that of -naphthaleneacetic acid. Immobilization of -decalylacetic acid is typically different. Bioassays showed -naphthaleneacetic acid as the sole biologically active component. It is concluded that stereo requirements necessary for biological activity are also required for polar auxin transport. It is further concluded that the observed specificity of the transport system is not related to the formation of immobilization products.     

Cytochemical and ultrastructural studies on normal and segregated nucleoli in meristematic cells

Summary The location of the two nucleolar components, in meristematic root tip cells ofAllium cepa has been studied. The segregation of granular and fibrous components of the nucleoli was induced by adenosine-3deoxyriboside. The relationship between the ultrastructure and the stain affinities of both nucleolar regions has been determined by using different selective nucleolar stains.The pars fibrosa appears positive in the silver, lead, zinc, and methylen blue stains and shows a high ATPase activity.     

Longer coleoptiles improve emergence through crop residues to increase seedling number and biomass in wheat (Triticum aestivum L.)

Crop residues protect soils from erosion, reduce soil water evaporation and increase soil organic matter. Yet management of stubbles for cropping can be difficult. Surface-retained residue can act as a mechanical barrier to slow emergence and reduce seedling biomass. Longer coleoptiles improve seedling emergence with deep sowing and may assist where stubble load is large. In a glasshouse study, six wheat and barley genotypes were sown at 30 and 50mm depth into pots containing pasteurised soil. Unweathered sorghum, canola and wheat stubble were added at 0, 3 and 6t/ha equivalents to the soil surface and pots watered above or below the stubble. Stubble species and watering regime had little effect on seedling growth. However, deeper sowing and increased stubble mass adversely affected most seedling characteristics particularly slowing seedling emergence and reducing tiller number to decrease plant biomass (environmental correlations (r_e) of –0.98** and 0.88**, respectively). Shorter coleoptile Rht-B1b wheats Banks and Janz, and barley Beecher emerged slower and abnormally with thicker stubble, and had more sterile tillers to reduce total tiller number and biomass. Deeper crowns for these genotypes also resulted in proportionally less biomass located above the stubble. The converse was true of long coleoptile Vigour 18, Halberd, and its Rht8 progeny, HM14bS which were less affected by stubble mass and sowing depth. In a corresponding field study, increasing wheat stubble mass from 0 to 3 and 6 t/ha delayed seedling emergence and decreased plant number to reduce biomass. Short coleoptile wheat genotypes Hartog and Janz emerged slower and produced less biomass at 3 and 6t/ha of stubble than long coleoptile wheat genotypes Halberd and HM14bS. Emergence of seedlings sown at 50mm depth with 6t/ha overlying stubble was similar to that sown at 120mm with no stubble, reflecting the similar impact of retained residues to deep sowing. Genetic variation for coleoptile length and availability of gibberellin-responsive dwarfing genes such as Rht8 will allow development of long coleoptile wheats for deep sowing or where stubble retention is practiced.     

Darwin’s Methodological Evolution

A necessary condition for having a revolution named after you is that you are an innovator in your field. I argue that if Charles Darwin meets this condition, it is as a philosopher and methodologist. In 1991, I made the case for Darwins innovative use of thought experiment in the Origin. Here I place this innovative practice in the context of Darwins methodological commitments, trace its origins back into Darwins notebooks, and pursue Darwins suggestion that it owes its inspiration to Charles Lyell.     

Role of local germplasm and induced mutations in the improvement of the protein content in rice

Summary Ninety local cultivars and 124 induced grain shape mutants were screened for their protein content and the distribution of protein in their endosperm. The protein content varied widely from 4.2 to 12.1% in the local varieties and from 5.5 to 13.7% in the mutants. Some high protein lines like Chinnavadlu, Budumavadlu, etc. with more than 10.0% protein were identified. Protein distribution studies have shown that several mutants, such as IR-8-FG-28, IR-8-FG-33, etc., have a deeper distribution of protein bodies and some local cultivars, like Kakathya and Muchulu, have a relatively more uniform distribution of protein in the endosperm. The protein content at different milling levels revealed that at 12% milling the percentage retention of protein varied from about 79 to 96% irrespective of the percentage of protein, suggesting that the distribution of protein, in addition to the protein content, plays an important role in nutritive value of rice.     

Characterization of the guanosine 3′∶5′-monophosphate-dependent protein kinase from silkworm eggs and analysis of the endogenous protein substrates

Summary In a previous paper, two types of cyclic nucleotide-dependent protein kinases, namely cGMP dependent G-kinase and cAMP dependent A-kinase, in silkworm eggs has been reported (Takahashi et al. 1975; Takahashi 1976). One of these, G-kinase, has now been purified 2400-fold by means of ammonium sulfate fractionation, chromatography on hydroxylapatite, DEAE cellulose, and gel filtration.Some of the properties of the enzyme are described. The enzyme is highly dependent on cGMP; it is strongly inhibited by GTP in a noncompetitive manner not only for ATP but also for cGMP. GTP was found to be highly inhibitory on G-kinases from various tissues of the silkworm, but did not inhibit the A-kinase.Incubation of the egg extract with [-³²P]ATP and Mg²⁺ led to the formation of three major³²P-labelled proteins, with molecular weights of 42.000, 70.000 and 180.000 as analyzed by SDS polyacrylamide gel electrophoresis. Two of them corresponded to the subunits of vitellin.The silkworm vitellin was effectively phosphorylated both by the highly purified G-kinase and by the A-kinase. It is concluded that the G-kinase is involved in the phosphorylation of vitellin in developing silkworm eggs.Abbreviations cAMP adenosine 35-monophosphate - cGMP guanosine 35-monophosphate - A-kinase adenosine 35-monophosphate-dependent protein kinase - G-kinase guanosine 35-monophosphate-dependent protein kinase - MIX 1-methyl-3-isobutylxanthine