Physical map of the bacteriophage L (Salmonella typhimurium)

Abstract A circular restriction map of the genome of the phage L ( Salmonella typhimurium ) has been constructed with five restriction endonucleases, Eca I, Eco RI, Bam HI, Bgl I, and Pst I. The Eco RI fragments of phage-L DNA were cloned into pACYC184, and the resulting recombinant plasmids pL1, pL2,…,pL7 were introduced into Salmonella typhimurium . The genes present on the fragments cloned were identified by the marker rescue experiments with the L-phage amber mutants. A physical gene map of the L genome obtained in this way was compared with that of P22.     

Studies on protein-phosphorylation reactions in isolated chromatin.

The endogenous protein-phosphorylating activity of isolated chromatin was tested. We have found that a group of high-molecular-weight proteins (Mr greater than 50 000) was preferentially phosphorylated when chromatin from mouse ascites cells or from bovine lymphocytes was incubated in the presence of ATP. After disintegration of chromatin by nuclease treatment or by high salt concentration, a larger spectrum of chromatin proteins becomes accessible for phosphorylation by the chromatin-bound protein kinase. Some observations described in this communication may help to partially explain this result. The protein kinase was not found in nucleosomal subunits, indicating a non-random distribution of the enzyme in chromatin. This suggests that enzyme and substrate have to be in close spatial contact for the phosphorylation reaction to occur. Furthermore, we have shown for one protein, histone H1, that phosphorylation sites for the endogenous protein kinase are available on the free but not on the DNA-bound protein, suggesting that phosphate-accepting sites in chromatin proteins may be blocked by protein-DNA or by protein-protein interactions. We also discuss the possibility that chromatin protein kinase occurs in stable complexes with its phosphate-accepting substrates, as has been suggested by the findings of other [Kish, V.M. & Kleinsmith, L.J. (1974) J. Biol. Chem. 249, 750-760].     

CHROMOSOME COUNTS OF COMPOSITAE FROM THE UNITED STATES AND MEXICO

Chromosome counts are reported for 126 taxa representing 122 species and 61 genera of Compositae. First reports include two genera, Stylocline (n = 14) and Chromolepis (n = 19), 17 species, two infraspecific taxa, and one interspecific hybrid. Five additional taxa have chromosome numbers differing from previously published accounts. Carminatia is reinstated to generic status.     

Chemical modifications of Achromobacter collagenase and their influence on the enzymic activity

A study of the influence of chemical modifications on the activity of Achromobacter iophagus collagenase (EC 3.4.24.8) has led to the following conclusions: a modification of 4 out of 80 COOH groups with carbodiimide led to 90% loss of enzymic activity. A 70% inactivation was found after modification of two tyrosines out of 30 with tetranitromethane. The modification of four to six tryptophans out of 16 with 2-hydroxy-5-nitrobenzyl bromide decreased enzyme activity to 36%. This inactivation is accelerated in the presence of collagen. An increase of reagent/enzyme molar ratio led to a modification of 16 tryptophan residues and denaturation of Acahromobacter collagenase. A modification of two arginines out of 18 with 1,2-cyclohexanedione and eight NH2 groups out of 24 with 2,3-dimethyl maleic anhydride does not change the collagenolytic activity. All NH2 groups become available for 2,3-dimethyl maleic anhydride after dissociation of the dimer. A possible analogy of hydrolytic site of collagenase with that of two other known bacterial metalloproteinases (thermolysin and Bacillus subtilis neutral proteinase (EC 3.4.24.4)) is discussed.     

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.     

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.