The syntheses and properties of novel, extremely strong uncharged polyaminophosphazene bases up to a high level of steric hindrance are described. Most of the systems were readily prepared in up to molar quantities and conveniently recovered from their salts. They are of appreciable to high chemical and thermal stability. Crystal structures of their salts were determined in order to parametrize a force field, which is utilized in molecular modeling studies. The latter offer a rationalization of the high conformational mobility of these systems. These bases cover a range of ca. 15 pK units in basicity and extend the range of uncharged auxiliary bases by ca. 19 pK units up to DMsopKsH+ values of 34-35. They are proposed as a novel class of auxiliary bases for deprotonation of very low acidic compounds where chemists have been so far left to classical metalorganic bases. Depending on the basicity range and the degree of steric protection of the basic center, these systems are particularly applicable to E2 elimination or to in situ generation of highly reactive "naked" anions.Uncharged nitrogen bases have a long tradition as widely used and often irreplaceable standard reagents in organic synthesis; many attempts to improve basicity and to reduce nucleophilicity have been reported"] since the classical work of Hunig et a1.L21. Until recently, amidines and guanidines as described by Eiter et al.f31, Eschenmoser et al.f4I, and Barton et al. ['] were generally considered the strongest synthetically useful auxiliary bases. In the early sixties there was a single report by Flynn et a1.L8] concerning applications of a somewhat stronger isobiguanide base. This base was even commercially available, but surprisingly has not been accepted by synthetic chemists.In connection with our own activities in this field we exploited among other s t r u~t u r e s [~~'~] the structural type of peralkylated triaminoiminophosphoranes. The simplest representative 1 was already known" l] and in our hands turned out to be of unprecedented base strength among kinetically active uncharged bases. The derivatives of this leading structure we synthesized s~bsequently['~,'~] proved to be chemically very stable, highly versatile and easy to recover auxiliary bases with a very broad range of steric shielding of the basic enter[',^*'^]. The Concept of Phosphazene BasesAt the outset there was the question, whether it would be possible to further enhance basicity by the same formal "homologization" which converts weakly basic tertiary amines to strongly basic triaminoiminophosphoranes.
Human tryptase, a mast-cell-specific serine proteinase that may be involved in causing asthma and other allergic and inflammatory disorders, is unique in two respects: it is enzymatically active only as a heparin-stabilized tetramer, and it is resistant to all known endogenous proteinase inhibitors. The 3-A crystal structure of human beta-tryptase in a complex with 4-amidinophenyl pyruvic acid shows four quasi-equivalent monomers arranged in a square flat ring of pseudo 222 symmetry. Each monomer contacts its neighbours at two different interfaces through six loop segments. These loops are located around the active site of beta-tryptase and differ considerably in length and conformation from loops of other trypsin-like proteinases. The four active centres of the tetramer are directed towards an oval central pore, restricting access for macromolecular substrates and enzyme inhibitors. Heparin chains might stabilize the complex by binding to an elongated patch of positively charged residues spanning two adjacent monomers. The nature of this unique tetrameric architecture explains many of tryptase's biochemical properties and provides a basis for the rational design of monofunctional and bifunctional tryptase inhibitors.
Thermus thermophilus HB27 is an extremely thermophilic, halotolerant bacterium, which was originally isolated from a natural thermal environment in Japan. This organism has considerable biotechnological potential; many thermostable proteins isolated from members of the genus Thermus are indispensable in research and in industrial applications. We present here the complete genome sequence of T. thermophilus HB27, the first for the genus Thermus. The genome consists of a 1,894,877 base pair chromosome and a 232,605 base pair megaplasmid, designated pTT27. The 2,218 identified putative genes were compared to those of the closest relative sequenced so far, the mesophilic bacterium Deinococcus radiodurans. Both organisms share a similar set of proteins, although their genomes lack extensive synteny. Many new genes of potential interest for biotechnological applications were found in T. thermophilus HB27. Candidates include various proteases and key enzymes of other fundamental biological processes such as DNA replication, DNA repair and RNA maturation.
Proteinase inhibitors are important negative regulators of proteinase action in vivo. We have succeeded in isolating two previously unknown polypeptides (HF6478 and HF7665) from human blood filtrate that are parts of a larger precursor protein containing two typical Kazaltype serine proteinase inhibitor motifs. The entire precursor protein, as deduced from the nucleotide sequence of the cloned cDNA, exhibits 15 potential inhibitory domains, including the Kazal-type domains, HF6478, HF7665, and 11 additional similar domains. An inhibitory effect of HF7665 on trypsin activity is demonstrated. Because all of the 13 HF6478-and HF7665-related domains share partial homology to the typical Kazal-type domain but lack one of the three conserved disulfide bonds, they may represent a novel type of serine proteinase inhibitor. The gene encoding the multidomain proteinase inhibitor, which we have termed LEKTI, was localized on human chromosome 5q31-32. As shown by reverse transcriptase-polymerase chain reaction and Northern blot analysis, it is expressed in the thymus, vaginal epithelium, Bartholin's glands, oral mucosa, tonsils, and the parathyroid glands. From these results, we assume that LEKTI may play a role in antiinflammatory and/or antimicrobial protection of mucous epithelia.Proteinases are enzymes required for nonspecific processes of digestion and intracellular protein turnover as well as specific proteolytic activation of inactive precursors of many regulatory proteins, such as enzymes and peptide hormones. In addition, they are involved in several processes of extracellular matrix remodeling. Depending on the nature of their reactive center, they are subdivided into the classes of serine, cysteine, aspartate, and metalloproteinases (for review see Ref. 1). To control the action of proteinases in vivo, organisms produce another group of proteins, namely the proteinase inhibitors (for review see Refs. 2-4). Indeed, many pathological effects are due to the non-regulated action of endogenously produced proteinases or such proteinases encoded or synthesized by viruses, bacteria, and parasites (for review see Ref. 5). For instance, a genetically determined fault of the ␣ 1 -proteinase inhibitor may lead to an enhanced proneness to lung emphysema caused by uncontrolled action of leukocyte elastase (6 -8). Thus, proteinase inhibitors represent an important therapeutic tool for a large number of different disorders.Here we report the isolation of two peptides (HF6478 and HF7665) from human blood filtrate (hemofiltrate), which may represent a novel class of proteinase inhibitor. Blood filtrate, a by-product of ultrafiltration of the blood from patients with acute renal failure, is routinely used by us as a source for the systematic as well as random isolation of novel human peptides (9). Due to the cut-off limit of the hemofilters (approximately 20,000 Da), it mainly contains peptides exhibiting a molecular mass below 20,000 Da. Nevertheless, we succeeded in isolating members of many different peptide/protein families suc...
Intraclonal genome diversity of Pseudomonas aeruginosa was studied in one of the most diverse mosaic regions of the P. aeruginosa chromosome. The ca. 110-kb large hypervariable region located near the lipH gene in two members of the predominant P. aeruginosa clone C, strain C and strain SG17M, was sequenced. In both strains the region consists of an individual strain-specific gene island of 111 (strain C) or 106 (SG17M) open reading frames (ORFs) and of a 7-kb stretch of clone C-specific sequence of 9 ORFs. The gene islands are integrated into conserved tRNAGly genes and have a bipartite structure. The first part adjacent to the tRNA gene consists of strain-specific ORFs encoding metabolic functions and transporters, the majority of which have homologs of known function in other eubacteria, such as hemophores, cytochrome c biosynthesis, or mercury resistance. The second part is made up mostly of ORFs of yet-unknown function. Forty-seven of these ORFs are mutual homologs with a pairwise amino acid sequence identity of 35 to 88% and are arranged in the same order in the two gene islands. We hypothesize that this novel type of gene island derives from mobile elements which, upon integration, endow the recipient with strain-specific metabolic properties, thus possibly conferring on it a selective advantage in its specific habitat.Genetic variability within bacterial species can be the result of nucleotide substitutions, intragenomic reshuffling, and acquisition of DNA sequences from another organism (3). The considerable impact of the last strategy, termed horizontal gene transfer, on microbial evolution and its integral role in the diversification and speciation of the bacteria has become apparent from recent analyses based on the growing pool of genomic sequence information (7,18,23,28). Prominent examples are the pathogenicity islands of many obligatory pathogens (14). These chromosomally encoded regions typically contain large clusters of virulence genes not present in closely related nonpathogenic strains and can, upon integration, transform a benign organism into a pathogen. Whereas the molecular mechanism of chromosomal integration has been resolved for some conjugative transposons and bacteriophages and details about the transmissibility of conjugative plasmids are well known, the evolution and mobility of gene islands remain obscure (14). Often these DNA blocks are integrated adjacent to or within tRNA genes, and some contain a phage-related integrase gene near one end, suggesting that gene islands may have been generated by a phage or by a plasmid with integrative functions (14, 42). Nevertheless, the comparative sequence analysis of gene islands so far have not pointed to any common genetic repertoire that confers transmission and acquisition.The gram-negative bacterium Pseudomonas aeruginosa is ubiquitously distributed in aquatic and soil habitats, and it is an opportunistic pathogen for plants, animals, and humans (38). No correlation between certain P. aeruginosa clones and disease habitats or environm...
The crystal structure of the complex formed between eglin c, an elastase inhibitor from the medical leech, and subtilisin Carlsberg has been determined at 1.2 A resolution by a combination of Patterson search methods and isomorphous replacement techniques. The structure has been refined to a crystallographic R‐value of 0.18 (8‐1.2 A). Eglin consists of a four‐stranded beta‐sheet with an alpha‐helical segment and the protease‐binding loop fixed on opposite sides. This loop, which contains the reactive site Leu45I‐‐Asp46I, is mainly held in its conformation by unique electrostatic/hydrogen bond interactions of Thr44I and Asp46I with the side chains of Arg53I and Arg51I which protrude from the hydrophobic core of the molecule. The conformation around the reactive site is similar to that found in other proteinase inhibitors. The nine residues of the binding loop Gly40I‐‐Arg48I are involved in direct contacts with subtilisin. In this interaction, eglin segment Pro42I‐‐Thr44I forms a three‐stranded anti‐parallel beta‐sheet with subtilisin segments Gly100‐‐Gly102 and Ser125‐‐Gly127. The reactive site peptide bond of eglin is intact, and Ser221 OG of the enzyme is 2.81 A apart from the carbonyl carbon.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
334 Leonard St
Brooklyn, NY 11211
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.