Enzymes as molecular automata: a reflection on some numerical and philosophical aspects of the hypothesis.

Enzymes, by means of their properties of specific recognition and allosteric modulation, are able to integrate many separate processes into systemic units with coherent functions; in a sense, they have to be considered as the true organizers of the cytoplasmic processes. In this respect, the present article describes a simple model, based on binary variables and automata theory, which simulates the basic regulatory performance of the modulated enzyme. The model admits a variety of modifications and improvements; it also suggests some original lines of thought on which to reflect about the organization and collective phenomena of the networks of enzymes. In discussing the connection of this 'molecular automata' hypothesis with other areas of present-day theoretical biology, a fertile panorama of initiatives appear. A special partnership between Information Science (computation) and Biology is developing.     

Enzymic memory. Steady state kinetic and physical studies with ascorbate oxidase and aspartate aminotransferase.

Enzymic memory is a kinetic phenomenon observable in double displacement mechanisms. The defining feature of enzymic memory is the occurrence of different rates of transfer for a common transferable group from the substituted enzymes obtained with different donor substrates. Memory behavior was previously demonstrated for both the bovine and human liver rhodaneses (EC 2.8.1.1). Steady state kinetic tests for enzymic memory have now been done with ascorbate oxidase (EC 1.10.3.3) and aspartate aminotransferase (EC 2.6.1.1). The results were positive with ascorbate oxidase, which showed an oxygen reactivity ratio of 1:20:300 for the reduced enzymes obtained with reductate, araboascorbate, and ascorbate, respectively. Results were negative for the aminotransferase tested with the alternate donors glutamate and cysteine sulfinate, with oxaloacetate as the common acceptor. The structural basis of the ascorbate oxidase results was probed by comparison of both the ultraviolet absorption and fluorescence spectra of the oxidized enzyme with those of the reduced forms obtained with ascorbate and reductate. The results are consistent with a conformational basis for the memory phenomenon.     

The sulfur-cyanolysis sites of serum albumin: metabolite competition studies

In the presence of a source of sulfane sulfur, a cyanolysis reaction catalyzed by serum albumin may contribute to cyanide detoxication. The active site for this catalysis by serum albumin has been investigated in competition studies with ligands that have known albumin binding sites. Despite complications caused by the occurrence of multiple primary and secondary sites for many ligands, the results show that the primary sites for bilirubin, steroids, indoles, aspirin, and palmitate are distinct from that for sulfur. Laurate is a tight-binding partial inhibitor of the cyanolysis reaction, competitive with cyanide rather than with sulfur. In view of the formal mechanism previously established for the catalyzed reaction, this result indicates that the sulfur-cyanolysis site is probably near the site occupied by laurate.     

Isolation and characterization of a prokaryotic sulfurtransferase

A sulfurtransferase has been purified to apparent homogeneity from the prokaryote Acinetobacter calcoaceticus lwoffi by conventional protein fractionation techniques. Steady-state kinetic studies of the enzyme revealed that its formal mechanism varies with the acceptor substrate employed. With inorganic thiosulfate as the sulfane sulfur-donor substrate and cyanide anion as the acceptor, the enzyme was shown to catalyze the reaction by a double displacement mechanism like that of mammalian rhodanese (thiosulfate:cyanide sulfurtransferase, EC 2.8.1.1). In contrast, with a thiol as the acceptor substrate at relatively high concentrations, the reaction proceeds by a single displacement mechanism, reminiscent of catalysis by another sulfur-transferase, thiosulfate reductase, glutathione-dependent (EC 2.8.1.3). When dithiothreitol is the acceptor substrate, the enzyme cycles through both the single and double displacement pathways, with the flux through each depending differentially on the concentration of dithiothreitol employed. In view of both the relaxed acceptor substrate specificity and the corresponding variability of formal mechanism, the more general name of sulfane sulfurtransferase is proposed for this bacterial enzyme.     

Effect of hyaluronidase treatment of intact cells on hyaluronate synthetase activity

Hyaluronidase treatment of mouse oligodendroglioma cells in monolayer culture resulted in a 4-5-fold stimulation of hyaluronate synthetase, assayed in washed membrane preparations [Philipson, L., & Schwartz, N. B. (1984) J. Biol. Chem. 259, 5017-5023]. We now report studies on the mechanism of the hyaluronidase-induced increase in the specific activity of the membrane-bound synthetase complex. The stimulation was dependent on the concentration of hyaluronidase but not on the particular bond cleaved or the nature of the product generated. Analysis of chain growth during cell-free synthesis by the disaccharide ratio method suggested that substantial internal labeling of hyaluronate chains had occurred. With both treated and untreated membranes, greater than 90% of incorporated (and recovered) radioactivity appeared in unsaturated disaccharides. Further analysis showed that hyaluronidase treatment increased both the rate of elongation and the rate of release of elongated chains from the enzyme complex. Hyaluronidase treatment also caused a change in the apparent steady-state kinetic patterns of double-reciprocal plots from intersecting lines for membranes from control cells to a family of parallel lines. Both the overall stimulation of synthesis and the change in apparent kinetic pattern were reversed by brief incubation of washed cells in the absence of hyaluronidase. These results have led to the development of an explicit kinetic model for hyaluronate synthesis which suggests an explanation for the switch in apparent kinetic patterns based on changing concentrations of a postulated key intermediate.     

The human genome contains multiple sequences of varying homology to mouse mammary tumour virus DNA

Sequences related to the mouse mammary tumour virus (MuMTV) DNA were isolated from a genomic library of human DNA by screening under conditions of relaxed stringency. It is estimated that there are in the order of 50 MuMTV-like sequences per haploid genome and that the homology between the different human sequences and MuMTV varies by 15%.     

An estrogen receptor from xenopus laevis liver possibly connected with vitellogenin synthesis

This paper describes an estrogen receptor which is found in both the nucleus and cytoplasm of liver cells from male Xenopus laevis, and which seems to be involved in the induction of vitellogenin synthesis. It has a high affinity for estradiol (K_d = 0.5 × 10^?9 M), and the affinities of various steroids for the receptor correlate well with their ability to induce vitellogenin synthesis. It sediments at 3.5S at 0°C in 0.5 M KCI. The rate of sedimentation is unaffected by incubation at 20°C prior to centrifugation, but increases if the salt concentration is lowered to 0.1 M KCI or to zero. It has a Stokes radius of 2.6 nm and a molecular weight of approximately 40, 000. The receptor is present at very low levels compared to other steroid target tissues (50–100 fold less than chick oviduct). The cytoplasm of a single hepatocyte contains 92 ± 18 binding sites for estradiol, while each nucleus contains 99 ± 19 sites.