Growth of Streptomyces bambergiensis and glucose isomerase biosynthesis

Summary Growth and glucose isomerase biosynthesis in Streptomyces bambergiensis ATCC 13879 have been studied under different conditions. Some data concerning correlation between cultivation conditions and elemental analysis of the cells are also presented.     

Carbohydrate nutrition of claviceps purpurea for alkaloid production related to the osmolality of media

Summary The effects of most frequently used carbohydrates and osmolalities of media on the growth and productivity ofClaviceps purpurea in submerged cultures were investigated. The maximum alkaloid yield was obtained with 20% mannitol as well as with 10% mannitol when 2% NaCl was added to the medium and the osmolality about 1 osmol/kg H₂O was established at the end of fermentation. Increased or decreased osmolality of the medium had a negative influence on fungal growth and alkaloid production. Almost the same effect was observed with sucrose as with mannitol, whereas with glucose only maximal growth was achieved.     

Length of human constitutive heterochromatin in relation to chromosomal contraction

Summary Linear measurement of blocks of constitutive heterochromatin and the euchromatin portion 1q-h in three members of a family was used to study the dependence of the size of C blocks on the degree of chromosomal contraction. The results demonstrate that the size of heterochromatin portions decrease regularly with an increases of the degree of euchromatin contraction. The dependence was found to be linear, except for mitoses with an extremely high or low degree of contraction. The finding was used for the development of a new method of evaluation of constitutive heterochromatin.     

The T-Cell factor specific for poly(Tyr,Glu)-Poly(Pro)-Poly(Lys) is an I-Region gene product

The nature of a T-cell factor specific for poly(Tyr,Glu)-poly(Pro)-poly(Lys) [(T,G)-pro-L] was established in the present study. The activity of the (T,G)-Pro-L-specific factor was not removed by anti-mouse immunoglobulin Sepharose columns, suggesting that it is not a classical immunoglobulin. On the other hand, the factor lost its activity after passage through immunoadsorbents prepared with anti-H-2 sera raised against theH-2 haplotypes of the mouse strains in which the factor was prepared. Furthermore, this factor was adsorbed byI region-specific antisera but not by antisera directed against theI-J andI-C subregions as well as theK andD regions of theH-2 complex. Thus, the (T,G)-Pro-L-specific T-cell factor is most probably anI-A subregion gene product.     

Aminoacid composition and hydrophobicity index of mitochondrial polypeptides

The aminoacid composition of protein stained bands in polyacrylamide gels, after electrophoresis of proteins from inner mitochondrial membranes, was investigated hydrolyzing directly the gel slices. The Hydrophobicity Index of 18 prominent polypeptide bands was calculated after their aminoacid analysis. The polypeptides less related to the membrane have low hydrophobicity as inferred from their Hydrophobicity Indexes.     

Stoicheiometry of dicyclohexylcarbodiimide-ATPase interaction in mitochondria

1. The oligomeric dicyclohexylcarbodiimide (DCCD)-binding protein of mitochondrial ATPase was studied using (a) the relationship between [¹⁴C]DCCD binding and inhibition of ATPase activities and (b) the analysis of the kinetics of inhibition. 2. The [¹⁴C]DCCD binding to bovine heart mitochondria is linearly proportional to the inhibition of ATP hydrolysis up to a 50% decrease of the original activity resulting in 0.6 mol DCCD bound covalently to the specific inhibitory site (Hous?t?k, J., Svoboda, P., Kopecký, J., Kuz?ela, S?. and Drahota, Z. (1981) Biochim. Biophys. Acta 634, 331–339) per mol of the fully inhibited enzyme. 3. Kinetics of the inhibition of both the ATPase activity (heart and liver mitochondria) and ADP-stimulated respiration (liver) reveal that 1 mol DCCD per mol ATPase eliminates both the synthetic and the hydrolytic activities. It is inferred that the activity-binding correlation underestimates the number of DCCD-reactive sites. 4. The second-order rate constant of the DCCD-ATPase interaction (k) is inversely related to the concentration of membranes, indicating that DCCD reaches the inhibitory site by concentrating in the hydrophobic (phospholipid) environment. 5. At a given concentration of liver mitochondria, comparable k values are obtained both for the inhibition of ATP hydrolysis (k=5.35·10²M^?1·min^?1) and ADP-stimulated respiration (k=5.67·10²M^?1·min^?1). 6. It is concluded that both the synthetic and the hydrolytic functions of ATPase are inhibited via a common single DCCD-reactive site. This site is represented by one of the several polypeptide chains forming the oligomer of the DCCD-binding protein. The inhibitor-ATPase interaction does not exhibit cooperativity, indicating that the preferential reactivity towards DCCD is an inherent property of the inhibitory site.     

Protein-lipid interactions and differential scanning calorimetric studies of bacteriorhodopsin reconstituted lipid-water systems

Bacteriorhodopsin has been reconstituted at various molar concentrations into liposomes of dimyristoyl- and also of dipalmitoylphosphatidylcholine. Differential scanning calorimetry indicates that as the protein concentration within the lipid bilayer increases, the cooperativity of the lipid phase transition is reduced, i.e. the transition is broadened, while the midpoint transition temperature remains virtually unchanged. Freeze-fracture electron microscopy of our preparation shows, in agreement with previous data from other laboratories, that extensive protein aggregation occurs when the liposome is cooled below the T_c transition temperature of the lipid. Laser flash photolysis measurements of protein rotation of the bacteriorhodopsin show, especially in the case of protein-rich recombinants, that protein aggregates exist even above T_c. The perturbation caused by the presence of bacteriorhodopsin in the lipid bilayer is similar to that produced by other intrinsic proteins. The difficulty of correlating the observed calorimetric enthalpy data with a simple concept of a ‘boundary lipid layer’ based upon consideration of a single isolated protein is discussed in view of the occurrence of protein aggregates both above and below T_c. It is concluded that the reduction of enthalpy is related to the number of lipids which solvate the protein aggregates within the protein-lipid patches and are thereby removed from the cooperative melting and enthalpy of the remaining regions of pure lipid.     

The primary structure of the Fab fragment of protein KAU, a monoclonal immunoglobulin M cold agglutinin

The complete amino acid sequence of the Fab fragment of protein KAU, a human monoclonal cold agglutinin (IgMk) with anti-I activity, was determined. The light chain (L-chain) consists of 215 residues; the variable (V)L region belongs to the Hum/Kv325/kIIIb sub-subgroup that is preferentially selected in human IgM autoimmune response. The joining (J) region is encoded by the Jk4 gene, and the constant region (C)L domain expresses the km3 allotypic marker. The Fd fragment contains 232 amino acids, and 120 of them comprise the variable domain. The VH region corresponds to the VHIV subgroup and is closely related to the VHIV 2.1 gene isolated from genomic DNA expressed in peripheral blood of a healthy Caucasian. The complementary-determining region 1 has a unique amino acid (Asp) at position 31, and the complementary-determining region 3 codified by the diversity segment (D) gene, shows poor homology with other known D sequences. The joining segment with two unusual substitutions at the D-J junction is encoded by the JH4 gene. Thus, cold agglutinin KAU is an IgM, VkIIIb-Jk4-km3; VHIV-JH4-C mu.     

On the mechanism of bacteriorhodopsin solubilization by surfactants

Purple membrane bacteriorhodopsin can be easily solubilized by Triton X-100 and other detergents, but not by deoxycholate. In order to understand this behavior, we have examined the effects of a variety of surfactants. We show that detergents containing the cholane ring (cholate, taurocholate, 3[(3-cholamidopropyl)diethyl-ammonio]propanesulfonic acid...) are virtually unable to solubilize native bacteriorhodopsin. However, when the protein is reconstituted in dimyristoyl phosphatidylcholine and solubilization is assayed at a temperature such that bacteriorhodopsin is in the form of monomers, solubilization by cholane detergents does occur. We propose that steric factors prevent access of the rigid planar surfactant molecules to the hydrophobic protein regions. These are perhaps located in the monomer-monomer interface, whose solvation by surfactants is essential for solubilization to occur. We note that the capacity of some detergents to solubilize bacteriorhodopsin is always associated within the same range of surfactant concentrations with bleaching (partial or total) of the protein chromophore. The detergent-induced bleaching is at least partially reversible, suggesting that free retinal remains associated to some membrane components. While some surfactant molecules remain tightly bound to the membrane protein, cholane detergents can be completely removed from bacteriorhodopsin. Our results indicate that a structure-function relationship exists for detergents applied to the solubilization of bacteriorhodopsin.