Engineering of zinc finger and MHC motifs to locked-in tertiary folds

The assembly of helical and -sheet peptide blocks containing reactive chain ends results inhighly branched chain architectures (locked-in folds) mimicking native tertiary structures.This molecular kit strategy allows to bypass the protein folding problem in protein de novodesign and gives access to protein mimetics of high thermodynamic stability. The validity ofthis concept is exemplified for the design and synthesis of locked-in folds mimicking the zincfinger and MHC folding motifs.     

Structure de la glande nasale externe deTiliqua rugosa (Reptilia,Scincidae) et rapports avec sa fonction

Summary The scincid lizardTiliqua rugosa possesses a large external nasal gland which is located intraconchally. Highly ramified tubules, imbedded primarily in the periphery of the gland, unite to form collecting ducts which empty into a short excretory canal. The diameter of the tubules increases progressively from 30. at the distal extremity of the gland to over 200 at the level of the collecting ducts. The intraglandular portion of the excretory canal is often dilated to form an ampulla. The thickness of the epithelium increases from 12 at the level of the tubules to 25–30 in the excretory canal.The excretory canal is lined with an epidermal epithelium close to the point where it enters the vestibule. In all the rest of the gland the tubules are lined with two cell types: large, typical muco-serous cells and striated cells. At the distal end of the tubules the striated cells are narrow and poorly differentiated and alternate more-or-less regularly with the muco-serous cells. The relative proportion of these striated cells increases progressively, as does their size, as one moves proximally down the tubule. In the gland as a whole the striated cells are approximately twice as numerous as the muco-serous cells but, due to their smaller size, they occupy less than one third of the tubular volume.Electron microscopy of the striated cells ofTiliqua rugosa revealed the presence of extensive lateral interdigitations and expansions of the basal cytoplasmic membrane, anatomical specialisations which are normally indicative of active salt transport. These modifications are less marked however than in the external nasal glands of the lizardsLacerta muralis andVaranus griseus, which do not appear to function as salt glands. In addition there are few mitochondria present, although they are of large size. The combination of these ultrastructural features, plus the fact that the striated cells are intermixed with muco-serous cells in the tubules, makes it most unlikely that the external nasal gland ofTiliqua rugosa is capable of elaborating an hyperosmotic fluid. What is more, this has never been conclusively demonstrated in this species in physiological studies.The progressive specialisation of the striated cells from the distal to the proximal section of the tubules poses the problem of the origin and differentiation of this cell type.A review of results obtained from the study ofTiliqua rugosa and other species of lizards shows that the nature of the relationship between structure and function of the external nasal gland is far from clear. The existence of salt glands, capable of excreting hyperosmotic solutions, is invariably linked with the presence in the gland of well-developed striated segments composed almost entirely of cells possessing extensive interdigitations of the lateral membranes. Amongst terrestrial lizards, nasal salt glands are usually found in herbivorous species and they are primarily adapted to the extrarenal excretion of potassium ions. The problem for carnivorous species is more often that of an excess of sodium rather than potassium ions and with the possible exceptionAcanthodactylus species, functional nasal salt glands have not been demonstrated in terrestrial carnivores, despite the presence in some cases of well-developed striated segments in the gland having a similar structure to those found in herbivores. In humid regions, carnivorous lizards probably never require extrarenal excretory mechanism and in arid regions their survival is assured by their capacity to tolerate hypernatraemia when confronted with excessive salt loads. Salt glands capable of eliminating sodium ions to any extent have only been described in two littoral species, an herbivorous iguanid and a carnivorous varanid. Unfortunately the structure of their respective nasal glands has not yet been described and their further study would be desirable.     

Carbon dioxide and water vapor exchange over a <Emphasis Type="Italic">Miscanthus</Emphasis>-type grassland: Effects of development of the canopy

The eddy correlation technique was employed to measure net ecosystem carbon dioxide (CO₂) (NEE) and water vapor exchange (LE) over a C3/C4 co-occurring wet temperate Miscanthus-type grassland in the Kanto plain of Japan in the 1999 growing season. The maximal mean canopy height and maximal leaf area index were 1.0m and 5.5, respectively. The daily maximal LE was approximately 540Wm^–2. The maximum value of daily accumulative LE was 16.3MJday^–1. Daily variation of the decoupling factor () suggests that in the morning LE decoupled with the atmosphere, and the available energy was the major driving force for LE, whereas in the afternoon LE coupled strongly with the atmosphere, and the atmospheric evaporative demand played a critical role in LE. The decline in (from 0.8 to 0.5) with the growing season demonstrates that LE decoupled from the atmosphere in the later growth season. The peak NEE value was 57.4µmolCO₂m^–2s^–1 (the positive value signifies the canopy carbon gain was from the air). The maximal daily integrated NEE was 1.06molCO₂m^–2day^–1 observed during the peak growth stage. A rectangular hyperbolic model was used to describe the relation between daytime NEE and incident photosynthetic photon flux density (PPFD). The net ecosystem CO₂ was not light-saturated up to a PPFD level of 2000µmol m^–2s^–1. The initial slope estimated with the NEE–PPFD response model was approximately 0.042molCO₂mol^–1photon on average. The canopy light compensation point ranged from 210 to 430µmolm^–2s^–1 with an average of approximately 310µmolm^–2s^–1. Both the initial slope and the canopy light compensation point decreased as the canopy senesced. The switch in dominance from C3 to C4 plants played an important role in the canopy fluxes.     

“Maturation” of DNA duplexes

Reassociation of typical single-copy DNAs, like E. coli DNA, even when performed at relatively low temperatures, results in the formation of perfect duplexes with thermal stability very close to that of the native DNA. In contrast, duplexes of mouse repeated DNA as well as duplexes of Streptomyces DNA prepared under the same conditions, show a low thermal stability and undergo post-reassociation changes upon prolonged incubation. These changes, called maturation of the DNA duplexes, result in increasing of their thermal stability. Some of the factors affecting the rate of maturation are studied. The implication of the maturation process in reassociation analysis and in characterization of the heterogeneity of DNA is discussed.     

DNA polymorphisms in the ψβ1- and β-globin gene regions in asian macaques

DNA polymorphisms in the ₁--globin gene region in nine Asian macaques(Macaca fuscata, M. mulatta, M. nemestrina, M. cyclopis, M. fascicularis, M. arctoides, M. radiata, M. maura, andM. assamensis) were examined using several restriction endonucleases and the human ₁, IVS2, and IVS2 probes. TheBamHI site 3 to the -globin gene was polymorphic inM. fuscata andM. mulatta, while the HincII site and the EcoRI site in the ₁-globin gene region was highly polymorphic inM. fuscata andM. mulatta, respectively. These polymorphic sites also seem to be present in other Asian macaques. The present study of the polymorphism at theBamHI site 3 to the -globin gene in Asian macaques supports, at the nuclear DNA level, the idea that thefascicularis group includingM. fuscata, M. mulatta, M. cyclopis, andM. fascicularis is different from other Asian macaque groups.This study was supported in part by the Cooperation Research Program of the Primate Research Institute, Kyoto University.     

Bacillus stearothermophilus KP 1064 pullulan hydrolase

Summary A pullulan hydrolase of Bacillus stearothermophilus KP 1064 was purified homogeneously. The molecular weight, Stokes radius, sedimentation coefficient (s₂₀, w), extinction coefficient at 280 nm and pH 6.8, and isoelectric point were estimated as 115,000, 4.16 nm, 5.5 S, 1.92 cm²·mg^-1 and 4.4, respectively. The enzyme consisted of two identical subunits each comprising a methionine residue at the NH₂-terminus. The enzyme hydrolysed pullulan, amylopectin, soluble starch, amylose, -and -limit dextrins, - and -cyclodextrins, phenyl-d-maltoside, maltotriose, and maltopentaose. The main products from amylose and pullulan were maltose and panose, respectively. The substrate specificity, along with the pattern of products, suggested the assignment of the enzyme to a unique type of maltogenic -amylase (1,4-d-glucan glucanohydrolase, EC. 3.2.1.1).Presented at the Annual Meeting of the Agricultural Chemical Society of Japan, Sendai, 30 March 1983.     

Cloned β1,4N-acetylgalactosaminyltransferase: subcellular localization and formation of disulfide bonded species

Cloned human 1,4N-acetylgalactosaminyltransferase (GalNAcT) catalyses the synthesis of the glycosphingolipids GM2, GD2, and gangliotriosylceramide. To determine the subcellular location of this enzyme and whether it exists in intermolecular disulfide bonded species, we stably transfected Chinese hamster ovary (CHO) cells with three myc epitope-tagged forms of the GalNAcT gene: the native enzyme; the lumenal domain of GalNAcT fused to the cytoplasmic and transmembrane domains ofN-acetylglucosaminyltransferase I (GNT); and the transmembrane and lumenal domains of GalNAcT fused to the cytoplasmic domain of the Iip33 form of human invariant chain in order to retain the enzyme in the endoplasmic reticulum (ER). Immunoelectron microscopic analysis with anti-myc revealed that GalNAcT/myc was present throughout the Golgi stack, the GNT/GalNAcT/myc form was restricted primarily to the medial Golgi cisternae, and the Iip33/GalNAcT/myc form was restricted to the ER. Cells transfected with each of the three constructs contained high levels of GM2 synthase activityin vitro, but only the GalNAcT/myc form and the GNT/GalNAcT/myc forms were able to synthesize the GM2 productin vivo. The enzyme produced by all three constructs was present in the transfected cells in a disulfide bonded form having a molecular size consistent with that of a homodimer or higher aggregate.Abbreviations GSL glycosphingolipid(s) - CHO Chinese hamster ovary - GSL structures: GM2 GalNAc1,4(NeuAc2,3)Gal1,4GlcCer - GD2 GalNac1,4(NeuAc2,8NeuAc2,3)Gal1,4GlcCer - GM3 NeuAc2,3Gal1,4GlcCer - Gg₃ GalNAc1,4Gal1,4GlcCer - LacCer Gal1,4GlcCer - GlcCer glucosylceramide - PBS-BSA phosphate buffered saline pH 7.4 containing 1% bovine serum albumin - GalNAcT N-acetylgalactosaminyltransferase - GNT N-acetylglucosaminyltransferase I - Iip33 p33 form of human invariant chain - HPTLC high performance thin layer chromatography - PCR polymerase chain reaction - BFA Brefeldin A This paper is dedicated to Professor Sen-itiroh Hakomori on the occasion of his 65th birthday.     

Potential induced in a spherical cell by an intracellular point source and an extracellular point sink

Summary The potential is calculated for all time, inside and outside a spherical cell for a point source of current inside the cell and a point sink located a finite distance outside the cell. The source and sink are step functions in time. An eigenfunction expansion is obtained, valid for arbitrary =_m a/_i , where _i and _m are the conductivities inside the cell and in the membrane, respectively, a is the cell radius and the membrane thickness. For small , the eigenfunction expansion is expanded in powers of . The time dependence of the potential contains transients with two widely differing time constants =C_m a/_i, where C_m is the membrane surface capacitance, and _m=/. Closed-form expressions are obtained for the two leading terms, for small , after the rapid transient is over. The remaining time dependence is only in the potential inside the cell, and is a simple exponential increase, independent of position within the cell. It is found that the transmembrane potential is insensitive to the location of the extracellular sink at long times, but not at short times. The dependence of the potential on location of source, sink, and observer is studied for long times after the quick transients are over. A uniqueness theorem is derived for the solution to Laplace's equation for the membrane boundary condition.This work was supported in part by NSF Grant No. GB-24965. Dr. Peskoff is the recipient of NIH Special Research Fellowship No. 1F03 GM 55849-01. Mr. Ramirez is a Ford Foundation Pre-doctoral Fellow.     

Yeast PAPS reductase: properties and requirements of the purified enzyme

The enzymatic mechanism of sulphite formation in Saccharomyces cerevisiae was investigated using a purified 3-phosphoadenylsulphate (PAPS) reductase and thioredoxin. The functionally active protein (M_R 80–85 k) is represented by a dimer which reduces 3-phosphoadenylyl sulphate to adenosine-3,5-bisphosphate and free sulphite at a stoichiometry of 1:1. Reduced thioredoxin is required as cosubstrate. Examination of the reaction products showed that free anionic sulphite is formed with no evidence for bound-sulphite(s) as intermediate. V _max of the enriched enzyme was 4–7 nmol sulphite · min^-1 · mg^-1 using the homologous thioredoxin from yeast. The velocity of reaction decreased to 0.4 nmol sulphite · min^-1 · mg^-1 when heterologous thioredoxin (from Escherichia coli) was used instead. The K _m of homologous thioredoxin was 0.6 · 10^-6 M, for the heterologous cosubstrate it increased to 1.4 · 10^-6 M. The affinity for PAPS remained practically unaffected (K _{m PAPS}: 19 · 10^-6 M in the homologous, and 21 · 10^-6 M in the heterologous system). From the kinetic data it is concluded that the enzyme followed an ordered mechanism with thioredoxin as first substrate followed by PAPS as the second. Parallel lines in the reciprocal and a common intersect in the Hanes-plots for thioredoxin were seen as indication of a ping-pong (with respect to thioredoxin) uni-bi (with respect to PAPS) mechanism.Abbreviations APS adenylyl sulphate - DTE dithioerythritol - DTT dithiothreitol - HPLC high performance liquid chromatography - IEF isoelectric focusing - LSC liquid scintillation counting - 3,5-PAP adenosine-3,5-bisphosphate - PAPS 3-phosphoadenylyl sulphate - PEP phospho-(enol)pyruvate - SDS-PAGE sodium dodecyl sulphate polyacrylamide gel electrophoresis - Tris 2-amino-2-hydroxymethyl-1,3-propanediol     

Germ-free and barrier-raised TGFβ1-deficient mice have similar inflammatory lesions

Barrier-raised transforming growth factor 1 (TGF1)-deficient mice consistently die before 35 days of age of a severe multiorgan inflammatory disease that can affect the skeletal muscle, heart, liver, pancreas, salivary gland, lung, oesophagus and stomach. The underlying cause of this disease is not known. To determine whether abnormal responsiveness of the immune system to the presence of enteric flora plays a causative role, a colony of TGF1-deficient and wild-type mice were raised in a sterile environment. Seven germ-free TGF1-deficient and 5 germ-free TGF1 wild-type mice were examined. Lesion development was analysed and compared with historical data on 50 barrier-raised TGF1 mutant mice and 32 barrier-raised wild-type mice. All germ-free TGF1-deficient mice died shortly after weaning, as do their barrier-raised counterparts. There was a significant delay in death in germ-free TGF1-deficient mice compared with barrier-raised mutant mice. However, there was no difference in the type, severity or incidence of lesions between TGF1 mutant mice raised under germ-free or barrier conditions. Germ- free wild-type mice had no lesions. It is concluded that microorganisms play a minimal role in disease induction in TGF1-deficient mice     

Rapid 13C/12C turnover during growth of brown shrimp (Penaeus aztecus)

Summary Using natural-abundance ¹³C/¹²C ratios as tracers, carbon turnover rates were determined for postlarval brown shrimp, Penaeus aztecus, in five laboratory growth experiments. Although tissue turnover in adult animals generally occurs during maintenance metabolism and is a function of time, turnover for young postlarval shrimp was accelerated during growth, and was primarily a function of weight gained rather than time. Metabolic loss of tissue carbon during growth was usually approximated by the function, Fraction lost=1-(initial weight/final weight). For shrimp that switch diets in the sea, model calculations show that this high turnover rate coupled with a four-fold weight increase suffices for shrimp to achieve a close isotopic resemblance of 1 or less (¹³C units) to the new diet.In accordance with these predictive calculations, shrimp which had increased in weight by a factor of four or more in the culture experiments showed essentially constant isotopic values reflecting their new diets. For these larger animals, the average animal-diet difference varied across three diets from-0.9 to +11, and the ¹³C range among individuals was 1.4 in each experiment.     

Pesticides and heavy metals in Danish streambed sediment

The role of streambed sediment as a sink for pesticides and heavy metals was investigated in 30 Danish lowland streams. The investigated streams drain catchments varying in hydrology, topography, soil type and land use. The <250 m newly accumulated fraction of the uppermost 1–2 cm layer of streambed sediment was analysed for 19 old and modern pesticides and 9 heavy metals. DDE was present in the sediment of all the streams. Of the herbicides, fungicides and insecticides currently in use, the most frequently detected was diuron (50.0%), fenpropimorph (66.7%) and lambda-cyhalothrin (6.7%), respectively. The pesticides detected in the highest concentration were fenpropimorph (1700 ng g^–1), propiconazole (130 ng g^–1) and isoproturon (110 ng g^–1). The heavy metals are listed in order of increasing median concentration: Cd (0.80 g g^–1), Co (9.1 g g^–1), As (12.0 g g^–1), Ni (19.0 g g^–1), Cr (19.2 g g^–1), Pb (19.7 g g^–1), Cu (20.1 g g^–1), V (28.5 g g^–1), Zn (103 g g^–1). The average number of pesticides detected in the 27 streams draining predominantly agricultural catchments was (3.7±2.0) being higher (p=0.077) than in the three streams draining non-agricultural catchments (1.7±0.6). Pesticides were significantly related to catchment size, soil type and hydrological regime. Several heavy metals (Cr, Cu, Pb, V and Zn) were related to urban activity and soil type.     

Antibacterial activity of Δ9-tetrahydrocannabinol and cannabidiol

The minimum inhibiting concentrations (MIC) of ⁹-tetrahydrocannabinol (THC) and cannabidiol (CBD) for staphylococci and streptococci in broth are in the range of 1–5 g/ml. In the same range, both compounds are also bactericidal. In media containing 4% serum or 5% blood the antibacterial activity is strongly reduced (MIC 50g/ml). Gram-negative bacteria are resistant to THC and CBD.     

Derivation of 13C chemical shift surfaces for the anomeric carbons of oligosaccharides and glycopeptides using ab initio methodology

The dependence between the anomeric carbon chemical shift and the glycosidic bond , dihedral angles in oligosaccharide and glycopeptide model compounds was studied by Gauge-Including Atomic Orbital (GIAO) ab initio calculations. Complete chemical shift surfaces versus and for d-Glcp-d-Glcp disaccharides with (11), (12), (13), and (14) linkages in both - and -configurations were computed using a 3-21G basis set, and scaled to reference results from calculations at the 6-311G^** level of theory. Similar surfaces were obtained for GlcNAcThr and GlcNAcSer model glycopeptides in - and -configurations, using in this case different conformations for the peptide moiety. The results obtained for both families of model compounds are discussed. We also present the determination of empirical formulas of the form ¹³C=f(,) obtained by fitting the raw ab initio data to trigonometric series expansions suitable for use in molecular mechanics and dynamics simulations. Our investigations are consistent with experimental observations and earlier calculations performed on smaller glycosidic bond models, and show the applicability of chemical shift surfaces in the study of the conformational behavior of oligosaccharides and glycopeptides.     

A relationship between plasmid structure, structural lability, and sensitivity to site-specific endonucleases in Neisseria gonorrhoeae

Summary Nearly all gonococcal strains carry a small phenotypically cryptic plasmid of approximately 4,200 basepairs. A detailed physical map of this plasmid has been constructed, revealing the presence of numerous putative inverted repeats. These studies also revealed the presence on the plasmid of recognition sequences for several site-specific endonucleases (particularly HpaII, MspI and AluI) that are particularly resistant to cleavage, and confirmed previous reports of structural lability. Both the sites that are resistant to cleavage, and the observed structural variation are associated with the inverted repetitive sequences.     

C(alpha) chemical shift tensors in helical peptides by dipolar-modulated chemical shift recoupling NMR

The C chemical shift tensors of proteins contain information on the backbone conformation. We have determined the magnitude and orientation of the C chemical shift tensors of two peptides with -helical torsion angles: the Ala residue in G*AL (=–65.7°, =–40°), and the Val residue in GG*V (=–81.5°, =–50.7°). The magnitude of the tensors was determined from quasi-static powder patterns recoupled under magic-angle spinning, while the orientation of the tensors was extracted from C–H and C–N dipolar modulated powder patterns. The helical Ala C chemical shift tensor has a span of 36 ppm and an asymmetry parameter of 0.89. Its ₁₁ axis is 116° ± 5° from the C–H bond while the ₂₂ axis is 40° ± 5° from the C–N bond. The Val tensor has an anisotropic span of 25 ppm and an asymmetry parameter of 0.33, both much smaller than the values for -sheet Val found recently (Yao and Hong, 2002). The Val ₃₃ axis is tilted by 115° ± 5° from the C–H bond and 98° ± 5° from the C–N bond. These represent the first completely experimentally determined C chemical shift tensors of helical peptides. Using an icosahedral representation, we compared the experimental chemical shift tensors with quantum chemical calculations and found overall good agreement. These solid-state chemical shift tensors confirm the observation from cross-correlated relaxation experiments that the projection of the C chemical shift tensor onto the C–H bond is much smaller in -helices than in -sheets.     

Heritable variation in the phaseolin protein of nondomesticated common bean,Phaseolus vulgaris L.

Summary Crude protein extracts from single seeds of nondomesticated Mexican bean accessions were analysed by SDS polyacrylamide gel electrophoresis for variability in phaseolin protein. Six new phaseolin types; M1, M2, M3, M4, M5, M6, which contained polypeptides within the same range of molecular weights (51,000 to 45,000 daltons) as occur in the S, T and C phaseolin types of cultivated beans were identified. No T and C types were found among the non-domesticated Mexican accessions, and the S type occurred in less than 7% of the seeds screened. Genetic analyses of F₂ progenies from crosses between Sanilac (S), and five of the M types showed that each M phaseolin phenotype was allelic to the S type and expressed codominantly.     

Adenosine-5′-phosphosulfate (APS) as sulfate donor for assimilatory sulfate reduction in Rhodospirillum rubrum

Crude extracts of Rhodospirillum rubrum catalyzed the formation of acid-volatile radioactivity from (³⁵S) sulfate, (³⁵S) adenosine-5-phosphosulfate, and (³⁵S) 3-phosphoadenosine-5-phosphosulfate. An enzyme fraction similar to APS-sulfotransferases from plant sources was purified 228-fold from Rhodospirillum rubrum. It is suggested here that this enzyme is specific for adenosine-5-phosphosulfate, because the purified enzyme fraction metabolized adenosine-5-phosphosulfate, however, only at a rate of 1/10 of that with adenosine-5-phosphosulfate. Further, the reaction with 3-phosphoadenosine-5-phosphosulfate was inhibited with 3-phosphoadenosine-5-phosphate whereas this nucleotide had no effect on the reaction with adenosine-5-phosphosulfate. For this activity with adenosine-5-phosphosulfate the name APS-sulfotransferase is suggested. This APS-sulfotransferase needs thiols for activity; good rates were obtained with either dithioerythritol or reduced glutathione; other thiols like cysteine, 2-3-dimercaptopropanol or mercaptoethanol are less effective. The electron donor methylviologen did not catalyze this reaction. The pH-optimum was about 9.0; the apparent K _m for adenosine-5-phosphosulfate was determined to be 0.05 mM with this so far purified enzyme fraction. Enzyme activity was increased with K₂SO₄ and Na₂SO₄ and was inhibited by 5-AMP. These properties are similar to assimilatory APS-sulfotransferases from spinach and Chlorella.Abbreviations APS adenosine-5-phosphosulfate - PAPS 3-phosphoadenosine-5-phosphosulfate - 5-AMP adenosine-5-monophosphate - 3-AMP adenosine-3-monophosphate - 3-5-ADP 3-phosphoadenosine-5-phosphate (PAP) - DTE dithiorythritol - GSH reduced glutathione - BAL 2-3-dimercaptopropanol     

Isoform-Specific Membrane Translocation of Protein Kinase C After Ischemic Preconditioning

Mild cerebral anoxic/ischemic/stress insults promote tolerance and thereby protect the brain from subsequent lethal anoxic/ischemic insults. We examined whether specific activation of PKC , , , or isoforms is associated with ischemic preconditioning (IPC) in rat brain. IPC was produced by a 2-minute global cerebral ischemia. Membrane and cytosolic fractions of the hippocampi were immunoblotted using specific antibodies for PKC, , , and . PKC showed a significant translocation to the membrane fraction from 30 min to 4 h and PKC at 4 h following IPC. In contrast, the membrane/cytosol ratio of PKC showed a tendency to decrease at 30 min and 8 h, and the membrane/cytosol ratio of PKC was significantly decreased from 30 min to 24 h following IPC. These findings indicate PKC isoform-specific membrane translocations in the hippocampus after brief global brain ischemia and suggest that activation of PKC and PKC may be associated with IPC-induced tolerance in the rat hippocampus.     

Phylogenetic relationship of zeins and coixins as determined by immunological cross-reactivity and Southern blot analysis

Zeins from Zea mays L cv. Maya and coixins from Coix lacryma-jobi L. cv. Adlay were fractionated to obtain -, -, and -zein and -, -, and -coixin. The -coixins were composed of 4 polypeptide classes of 27 kDa (C1), 25 kDa (C2), 17 kDa (C4) and 15 kDa (C5) with solubility properties very similar to those of the 22 kDa and 19 kDa -zeins. Like the -zeins, the C1 and C2 -coixins corresponded to 80% of total Coix prolamins. The fraction corresponding to -coixin contained only one protein band of 22 kDa (C3). This coixin fraction has solubility properties similar to those of -zein and represents 15% of the total coixin. The -zein fraction was composed of a major 17 kDa protein band, while the -coixin fraction consisted of a mixture of - and -coixins.Polyclonal antibodies raised against C1 recognized C1 and C2 and cross-reacted strongly with the 22 kDa -zein, as did C4 and C5 antisera. The antiserum against -coixin showed strong cross-reaction with -zein. The homology between coixins and zeins was further investigated by using Southern hybridization analyses. The genomic DNA of maize and Coix were digested with several restriction enzymes and probed with cDNA clones representing 19 and 22 kDa -zeins as well as the 28 and 16 kDa -zeins. The Coix genome showed complex cross-hybridization sequences with the 22 kDa -zein cDNA, while no cross-hybridization was observed with the 19 kDa cDNA clone. The cDNA clone representing the 28 kDa -zein cross-hybridized with only one band of Coix genomic DNA, in contrast to the three bands observed in maize. This same Coix sequence also cross-hybridized with the cDNA clone representing the 16 kDa -zein. The relevance of these findings are discussed in the context of the origin of zein and coixin genes.