Stimulation by interferon of induction of differentiation of human promyelocytic leukemia cells

F₁-ATPase was isolated from yeast $\text{S.}\text{cerevisiae}$. The constituent subunits 1 and 2 were purified by gel permeation chromatography, and their amino acid compositions determined. Both subunits have a similar composition except for $\text{1}\text{2}$ cystine, methionine, leucine, histidine, and tryptophan. When F₁ is treated for three hours with 5′-p-[³H]fluorosulfonylbenzoyl adenosine in dimethylsulfoxide, 90% of the activity is lost. Disc gel electrophoresis of the modified complex showed that over 90% of the label was associated with subunit 2. A labelled peptide from a $\text{S.}\text{aureus}$ digest of subunit 2 was isolated and sequenced. It had the following amino acid sequence: His-Try¹-Asp-Val-Ala-Ser-Lys-Val-Gln-Glu, whereby Tyr¹ is the modified amino acid residue. This sequence shows homology to other sequences obtained from maize, beef heart, and $\text{E.}\text{coli}$ F₁-ATPases.     

Possible involvement of aquaporins in water uptake by pea seeds differing in quality

Using the method of room temperature phosphorescence (RTP), we divided air-dry pea (Pisum sativum L.) seeds subjected to accelerated ageing (40°C, 85% relative humidity) into three fractions: (I) high-quality seeds, (II) weakened seeds, and (III) dead seeds. In the process of ageing, seed germinability firstly decreased and then increased due to so-called “improved” seeds of fraction II, which returned to fraction I as judged from the RTP level; the germinability of these seeds became equal to that of fraction I seeds. Seeds capable of germination (fractions I and II) differed in the rates of imbibition, which depended on plasma membrane permeability (opened or closed water channels) but not on the presence of the seed coat. A low activation energy of seed imbibition in fraction II (less than 5 kcal/mol) indicates that water channels are open. A mercury-containing compound (5 μM p-chloromercuribenzoate (PCMB) reduced the rate of water uptake by these seeds, and dithiothreitol restored it. A high activation energy of fraction I seed imbibition (more than 12 kcal/mol) corresponded to the water uptake mainly across the lipid bilayer when water channels are closed. PCMB did not affect the rate of fraction I seed imbibition. We supposed that mature air-dry pea seeds had open water channels. During the first stages of fraction I seed imbibition, these channels were closed, limiting water uptake. NaF (100 μM), an inhibitor of phosphatase, prevented channel closing and accelerated the imbibition of fraction I seeds. It did not affect the imbibition rate of fraction II seeds, indicating their water channels to be opened. However, NaF did not affect the water uptake of “improved” fraction II seeds as well. It seems likely that their channels were closed during accelerated ageing but otherwise than via dephosphorylation. The results obtained indicate the possibility of water inflow regulation in the weakened seeds via the state of aquaporins, which form water channels in the membranes.     

A unique proenkephalin-converting enzyme purified from bovine adrenal chromaffin granules

A unique proenkephalin converting enzyme specifically generating enkephalin was partially purified from lysates of adrenal chromaffin granules. The enzyme, whose molecular weight is estimated as ca. 220,000, is thiol-dependent protease, with optimal pH at around 5.5. The enzyme converts proenkephalin to enkephalins by cleaving specifically at the sites of consecutive basic amino acid residues. The enzyme also converts BAM-12P, an adrenal “big” Met-enkephalin, to Met-enkephalin in a similar manner. During the enzyme reaction, formation of [Arg⁶]-Met-enkephalin was not observed. Additionally, [Arg⁶]-enkephalins were not converted to enkephalins by the enzyme. Consequently, the enzyme was proved to be a unique converting enzyme distinct from either trypsin-like or carboxypeptidase B-like proteases.     

Evidence for essential thiol groups and disulfide bonds in agonist and antagonist binding to the dopamine receptor

Dithiothreitol (DTT), a disulfide reducing agent, diminished the specific binding of [³H] dopamine to partially purified calf striatal membranes (P₂) but did not have an effect on [³H] spiroperidol binding. The thiol reagents, p-chloromercuribenzoate (PCMB), N-ethylmaleimide (NEM) and iodoacetamide (IA), were also tested for inhibitory effects on agonist and antagonist binding to the dopamine receptor. PCMB inhibited both [³H] dopamine and [³H] spiroperidol binding by changing the affinity (K_d) and the number of binding sites (B_max) for both of these ligands. This effect of PCMB was reversed by the addition of DTT. NEM inhibited binding to the dopamine agonist site but not to the antagonist site, while IA was ineffective on either site. These results indicate that a DTT-reducible disulfide bond may be an essential component for agonist binding to the dopamine receptor. Furthermore, the experiments with PCMB, NEM and IA suggest that the exposure of thiol groups in the dopamine receptor may play an important role in agonist and antagonist binding.     

The activation of glucose dehydrogenase by p-chloromercuribenzoate

Summary P-Chloromercuribenzoate alters various reactions of rat liver glucose (hexose phosphate) dehydrogenase differently. The reagent has little effect on the glucose: NAD or the glucose: NADP oxidoreductases, doubles the rates of oxidations of galactose-6-phosphate and glucose-6-phosphate by NADP and greatly stimulates the oxidations of glucose-6-phosphate and galactose-6-phosphate by NAD. The reagent appears to react with a sulfhydryl group of the enzyme since activation is reversed and prevented by mercaptoethanol. The direct reaction of the reagent with the enzyme is indicated by its lower thermal stability in the presence of the p-chloromercuribenzoate. The size of the enzyme appears to be the same when determined by sucrose gradient centrifugation in the presence or absence of p-chloromercuribenzoate. In microsomes, the oxidation of NADH or NADPH hampers measurements of glucose dehydrogenase. Since p-chloromercuribenzoate inhibits microsomal oxidation of reduced nicontinamide nucleotides, it is possible to assay for glucose dehydrogenase accurately in the presence of the mercurial in microsomes and microsomal extracts and thus measure the effectiveness of a detergent in extracting the enzyme from microsomes.Abbreviation pcMB p-chloromercuribenzoic acid