Internal structure of the silk fibroin gene of Bombyx mori. I The fibroin gene consists of a homogeneous alternating array of repetitious crystalline and amorphous coding sequences

The DNA sequence orgainzation of the protein encoding region of the gene for silk fibroin has been analyzed. The accompanying paper (Manningm R. F., and Gage, L. P. (1980) J. Biol. Chem. 255, 9451-9457) shows that the total length of the gene, and its protein, as well as the pattern of restriction sites in the gene is highly polymorphic among inbred stocks of Bombyx mori, In this paper, those features of fibroin gene structure which are invariant among these alleles are presented. Fibroin is composed primarily of relatively short "crystalline" and "amorphous" peptides of known sequence whose arrangement in the protein is unknown. Knowledge of the codons most commonly used in fibroin mRNA allowed utilization of particular restriction inzymes as a means for determing the nature and organization of crystalline and amorphous coding sequences in the fibroin gene. Three restriction endonucleases were identified that cleve sequences coding for amorphous region peptides. Their cleavage pattern revelaed that the repetitive coding sequence of the gene core (approximately 15 kilobases) is divided into at least 10 large crystalline coding domains interrupted by smaller amorphous coding domains. Many restriction endoncleases do not cleave the fibroin core at all, three of them with four gase recognition sequences. Specific deductions as to codon usage and repetitive sequence homogeneity in the gene follow from these results. One novel finding is the rigorous exclusion of the glycine codon GGA prior to serine codons even though this glycine codon is used frequently prior to alanine codons. The sequence homogeneity and the regularly alternating arrangement of crystalline and amorphous coding sequences of the gene are discussed in terms of the function of fibroin protein and the evolution of highly repetitive DNA.     

On the application of polyelectrolyte limiting laws to the helix–coil transition of DNA. V. Ionic effects on renaturation kinetics

The bimolecular rate constant k₂ for the association of complementary polynucleotide strands has been observed to increase strongly with increasing ionic strength—in fact, proportional to its third or fourth power. This effect is here interpreted quantitatively by means of polyelectrolyte theory starting with the Wetmur–Davidson postulate of a pre-equilibrium between separated strands and aligned segments close to one another but unbonded. The correct form, a power dependence of k₂ on ionic strength, is predicted. Comparison of the theoretical exponent with data allows the conclusion that each of the two single-stranded segments in the aligned but unbonded configuration consists of about 13–16 nucletides (not to be confused with the much smaller number of bonded base pairs in the nucleus), and that this number, denoted by Q, is possibly correlated either with a minimum length for duplex stability or with the persistence length of a single polynucleotide strand. It is suggested that experimental determination of the dependence of Q on (G+C)-content may distinguish between these possibilities. It is also suggested that addition of sufficient amounts of divalent metal ions such as Mg²⁺, Ca²⁺, or Co²⁺ may reverse the dependence of k₂ on ionic strength; under these conditions, k₂ is predicted to decrease with about the first power of ionic strength. At fixed ionic strength, k₂ should increase with increasing concentration of divalent metal ion, and, in fact, the published observation that the formation of poly(A)·2 poly(U) from poly(A)·poly(U) and poly(U) is second order in Mg²⁺ concentration is here correctly predicted from a priori molecular considerations. Finally, published association rate data for oligonucleotides are discussed in the present theoretical context.     

A new method of in situ hybridization

A new method for gene mapping at the chromosome level using in situ hybridization and scanning electron microscopy is described and has been applied to mapping the rRNA genes of Drosophila melanogaster. Biotin is covalently attached to Drosophila rRNA via a cytochrome c bridge at a ratio of one cytochrome-biotin per 130 nucleotides by a chemical procedure. Polymethacrylate spheres with a diameter of ca. 60 nm are prepared by emulsion polymerization and are covalently attached to the protein avidin at a ratio of 5–20 avidins per sphere. The biotin-labeled rRNA is hybridized to denatured DNA in a chromosome squash. Upon incubation with a sphere solution, some of the biotin sites become labeled with spheres because of the strong non-covalent interaction between biotin and avidin. The chromosome squash is examined in the scanning electron microscope (SEM). Polymer spheres, which are visible in the SEM, are observed to label the nucleolus, where the rRNA genes are located.Contribution number 5121 from the Department of Chemistry.     

Reactivity of cyanate with valine-1 (alpha) of hemoglobin. A probe of conformational change and anion binding.

The 3-fold increase in the carbamylation rate of Val-1 (alpha) of hemoglobin upon deoxygenation described earlier is now shown to be a sensitive probe of conformational change. Thus, whereas this residue in methemoglobin A is carbamylated at the same rate as in liganded hemoglobin, upon addition of inositol hexaphosphate its carbamylation rate is enhanced 30% as much as the total change in the rate between the CO and deoxy states. For CO-hemoglobin Kansas in the presence of the organic phosphate, the relative increase in the carbamylation rate of this residue is about 50%. These results indicate that methemoglobin A and hemoglobin Kansas in the presence of inositol hexaphosphate do not assume a conformation identical with deoxyhemoglobin but rather form either a mixture of R and T states or an intermediate conformation in the region around Val-1 (alpha). Studies on the mechanism for the rate enhancement in deoxyhemoglobin suggest that the cyanate anion binds to groups in the vicinity of Val-1 (alpha) prior to proton transfer and carbamylation of this NH2-terminal residue. Thus, specific removal with carboxypeptidase B of Arg-141 (alpha), which is close to Val-1 (alpha) in deoxyhemoglobin, abolishes the enhancement in carbamylation. Chloride, which has the same valency as cyanate, is a better competitive inhibitor of the carbamylation of deoxyhemoglobin (Ki = 50 mM) compared with liganded hemoglobin. Nitrate and iodide are also effective inhibitors of the carbamylation of Val-1 (alpha) of deoxyhemoglobin (Ki = 35 mM); inorganic phosphate, sulfate, and fluoride are poor competitive inhibitors. The change in pKa of Val-1 (alpha) upon deoxygenation may be due to its differential interaction with chloride.     

Sexual reproduction and parent-offspring conflict in the RNA "Tumour" virus/host relationship: implications for vertebrate oncogene evolution.

Longmuir and co-workers have reported that respiration of certain tissue slices is approximated by Michaelis-Menten kinetics. From this and other experimental findings, Longmuir proposed that a carrier is involved in tissue oxygen transport. Gold developed a deterministic model to examine this hypothesis. This report presents a stochastic model for a fixed site carrier in a more general framework that includes the stochastic counter-part to Gold's deterministic model as a special case. The kinetics of tissue oxygen consumption predicted by the model are examined for various cases.     

The impact of interventions in the global land and agri‐food sectors on Nature’s Contributions to People and the UN Sustainable Development Goals

Interlocked challenges of climate change, biodiversity loss, and land degradation require transformative interventions in the land management and food production sectors to reduce carbon emissions, strengthen adaptive capacity, and increase food security. However, deciding which interventions to pursue and understanding their relative co‐benefits with and trade‐offs against different social and environmental goals have been difficult without comparisons across a range of possible actions. This study examined 40 different options, implemented through land management, value chains, or risk management, for their relative impacts across 18 Nature's Contributions to People (NCPs) and the 17 Sustainable Development Goals (SDGs). We find that a relatively small number of interventions show positive synergies with both SDGs and NCPs with no significant adverse trade‐offs; these include improved cropland management, improved grazing land management, improved livestock management, agroforestry, integrated water management, increased soil organic carbon content, reduced soil erosion, salinization, and compaction, fire management, reduced landslides and hazards, reduced pollution, reduced post‐harvest losses, improved energy use in food systems, and disaster risk management. Several interventions show potentially significant negative impacts on both SDGs and NCPs; these include bioenergy and bioenergy with carbon capture and storage, afforestation, and some risk sharing measures, like commercial crop insurance. Our results demonstrate that a better understanding of co‐benefits and trade‐offs of different policy approaches can help decision‐makers choose the more effective, or at the very minimum, more benign interventions for implementation.