The three-dimensional solution structure of the recombinant B domain (FB) of staphylococcal protein A, which specifically binds to the Fc portion of immunoglobulin G, was determined by NMR spectroscopy and hybrid distance geometry-dynamical simulated annealing calculations. On the basis of 692 experimental constraints including 587 distance constraints obtained from the nuclear Overhauser effect (NOE), 57 torsion angle (phi, chi 1) constraints, and 48 constraints associated with 24 hydrogen bonds, a total of 10 converged structures of FB were obtained. The atomic root mean square difference among the 10 converged structures is 0.52 +/- 0.10 A for the backbone atoms and 0.98 +/- 0.08 A for all heavy atoms (excluding the N-terminal segment from Thr1 to Glu9 and the C-terminal segment from Gln56 to Ala60, which are partially disordered). FB is composed of a bundle of three alpha-helices, i.e., helix I (Gln10-His19), helix II (Glu25-Asp37), and helix III (Ser42-Ala55). Helix II and helix III are antiparallel to each other, whereas the long axis of helix I is tilted at an angle of about 30 degrees with respect to those of helix II and helix III. Most of the hydrophobic residues of FB are buried in the interior of the bundle of the three helices. It is suggested that the buried hydrophobic residues form a hydrophobic core, contributing to the stability of FB.(ABSTRACT TRUNCATED AT 250 WORDS)

Newly synthesized benzamide derivatives were evaluated for their inhibitory activity against histone deacetylase. The structure of these derivatives was unrelated to the known inhibitors, and IC 50 values of the active compounds were in the range of 2-50 µM. Structure-activity relationship on the benzanilide moiety showed that the 2′-substituent, an amino or hydroxy group, was indispensable for inhibitory activity. Although the electronic influence of the substituent in the anilide moiety showed only a small effect on inhibitory activity, the steric factor in the anilide moiety, especially at positions 3′and 4′, played an important role in interaction with the enzyme. Among these benzamide derivatives, MS-275 (1), which showed significant antitumor activity in vivo, has been selected for further investigation.

Neurofibrillary tangles, composed of hyperphosphorylated tau fibrils, are a pathological hallmark of Alzheimer's disease; the neurofibrillary tangle load correlates strongly with clinical progression of the disease. A growing body of evidence indicates that tau oligomer formation precedes the appearance of neurofibrillary tangles and contributes to neuronal loss. Here we show that tau oligomer formation can be inhibited by compounds whose chemical backbone includes 1,2-dihydroxybenzene. Specifically, we demonstrate that 1,2-dihydroxybenzene-containing compounds bind to and cap cysteine residues of tau and prevent its aggregation by hindering interactions between tau molecules. Further, we show that orally administered DL-isoproterenol, an adrenergic receptor agonist whose skeleton includes 1,2-dihydroxybenzene and which penetrates the brain, reduces the levels of detergent-insoluble tau, neuronal loss and reverses neurofibrillary tangle-associated brain dysfunction. Thus, compounds that target the cysteine residues of tau may prove useful in halting the progression of Alzheimer's disease and other tauopathies.

ABSTRACTcDNA clones encoding chum salmon (Oncorhynchus keta) growth hormone (sGH) have been isolated from a cDNA library prepared from chum salmon pituitary gland poly(A)+ RNA. Synthetic oligodeoxynucleotide mixtures based on amino acid residues 23-28 of sGH were used as hybridization probes to select recombinant plasmids carrying the sGH coding sequence. The complete nucleotide sequence of sGH cDNA has been determined. The cDNA sequence codes for a polypeptide of 210 amino acids, including a putative signal sequence of 22 amino acids. The 5' and 3' untranslated regions of the message were 64 and 426 bases long, respectively. Mature sGH was efficiently expressed in Escherichia coli carrying a plasmid in which the sGH cDNA was under control of the E. coli trp promoter; sGH comprised about 15% of the total cellular protein in such bacteria. The partially purified sGH from E. coli stimulated the growth of rainbow trout and the activity was indistinguishable from that of natural sGH.Human growth hormone now can be produced by genetically engineered organisms and can be used as a therapeutic agent. Salmon growth hormone (sGH) can be synthesized by use of similar techniques, and the massive supply of sGH may be extremely important to fish culture. Growth hormone (GH), together with prolactin and chorionic somatomammotropin (placental lactogen), forms a set of proteins that are structurally related and have partially overlapping biological activities (1). Primary structure analysis of the peptides and of the genes suggests that these hormone genes evolved from a common ancestral origin (1-5). Therefore, these genes provide an excellent model system for studying structurefunction relationships, evolution, and regulation of expression. GH genes have been isolated from several mammalian species and characterized in detail (3,(6)(7)(8). To obtain information about the evolution and the mechanisms of organization of this set of genes, it is essential to compare the structures of these hormone genes isolated from many organisms at various evolutionary stages. No information, however, has been available about lower vertebrates such as fish.

Neurofilaments (also called 10-nm filaments or intermediate filaments) from bovine brain were incubated with microtubule protein at 37"C in the presence or absence of 1 mM ATP and in a buffer that allowed microtubule assembly. Fallingball viscometry revealed that the (non-Newtonian) apparent viscosity of the ATP-containing mixtures is 5-20 times greater than that of the mixtures prepared without ATP. A larger ATP-dependent increase in viscosity (approximately 100-fold) was seen when purified tubulin replaced microtubule protein. The (1,4,5), and occasional observations of highly ordered arrays of neurofilaments surrounding microtubules (6) suggest together that the two structures are connected in vivo as part of an axoplasmic filamentous network. Neurofilaments are transported in the slow component of axonal transport (7,8), and it has been suggested (7) Neurofilaments, Tubulin, and Microtubule-Associated Proteins (MAPs). Neurofilaments were prepared from bovine brain as described (17). Microtubule protein referred to as "threetimes-cycled" was prepared by three cycles of the assembly/ disassembly method of Shelanski and coworkers (14, 18). Phosphocellulose (PC)-purified tubulin was prepared from twicecycled microtubule protein (19,20). The fraction that was initially retained by the PC column was eluted in buffer A containing 0.8 M NaCl and centrifuged for 60 min at 95,500 X g to remove any neurofilaments (see ref. 15). This material is referred to as "whole MAPs. " Aliquots of the whole MAP fraction were heated to 100°C for 5 min and then were centrifuged at 4°C for 20 min at 25,000 X g to remove flocculated protein (21). This material is referred to as "heat-treated MAPs." Purified MAP2 was prepared as described by Kim et al. (22). Upon NaDodSO4/polyacrylamide gel electrophoresis by the method of Laemmli (23), each of these five preparations appeared quite similar in protein composition to those described in the references given. The tubulin polymerized in buffer A at protein concentrations greater than about 2.7 mg/ml. Each preparation was dialyzed against buffer A and then stored in liquid N2. Aliquots were thawed, centrifuged for 10 min in a clinical centrifuge at 4°C, and used within 6 hr.Viscometry. Falling-ball viscometers were constructed and employed exactly as described by . Protein solutions were mixed at 4°C, and appropriate additions were made of nucleotide triphosphate solutions or of equal volumes of buffer in the cases of the controls. The cold solutions were immediately drawn into the viscometers, which were then immersed in a Tamson water bath controlled to +0.010C at 37. 0°C. The mixing and loading procedure required Abbreviations: MAP(s), microtubule-associated protein(s); PC, phosphocellulose; AdoPP[NH]P, adenyl-5'-yl imidodiphosphate. t Present address: