Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An experimental approach to evaluate the net binding free energy of buried hydrogen bonds and salt bridges is presented. The approach, which involves a modified multiple-mutant cycle protocol, was applied to selected interactions between TEM-1-beta-lactamase and its protein inhibitor, BLIP. The selected interactions (two salt bridges and two hydrogen bonds) all involving BLIP-D49, define a distinct binding unit. The penta mutant, where all side-chains constructing the binding unit were mutated to Ala, was used as a reference state to which combinations of side-chains were introduced. At first, pairs of interacting residues were added allowing the determination of interaction energies in the absence of neighbors, using double mutant cycles. Addition of neighboring residues allowed the evaluation of their cooperative effects on the interaction. The two isolated salt bridges were either neutral or repulsive whereas the two hydrogen bonds contribute 0.3 kcal mol(-1 )each. Conversely, a double mutant cycle analysis of these interactions in their native environment showed that they all stabilize the complex by 1-1.5 kcal mol(-1). Examination of the effects of neighboring residues on each of the interactions revealed that the formation of a salt bridge triad, which involves two connected salt bridges, had a strong cooperative effect on stabilizing the complex independent of the presence or absence of additional neighbors. These results demonstrate the importance of forming net-works of buried salt bridges. We present theoretical electrostatic calculations which predict the observed mode of cooperativity, and suggest that the cooperative networking effect results from the favorable contribution of the protein to the interaction. Furthermore, a good correlation between calculated and experimentally determined interaction energies for the two salt bridges, and to a lesser extent for the two hydrogen bonds, is shown. The data analysis was performed on values of DeltaDeltaG(double dagger)K(d) which reflect the strength of short range interactions, while DeltaDeltaG(o)K(D) values which include the effects of long range electrostatic forces that alter specifically DeltaDeltaG(double dagger)k(a) were treated separately.
J Mol Biol 2000 May 05
PMID:Evaluation of direct and cooperative contributions towards the strength of buried hydrogen bonds and salt bridges. 1077 66

Mutagenesis studies and alignments of homologous sequences have demonstrated that protein function typically is compatible with a variety of amino-acid residues at most exterior non-active-site positions. These observations have led to the current view that functional constraints on sequence are minimal at these positions. Here, it is shown that this inference assumes that the set of acceptable residues at each position is independent of the overall sequence context. Two approaches are used to test this assumption. First, highly conservative replacements of exterior residues, none of which would cause significant functional disruption alone, are combined until roughly one in five have been changed. This is found to cause complete loss of function in vivo for two unrelated monomeric enzymes: barnase (a bacterial RNase) and TEM-1 beta-lactamase. Second, a set of hybrid sequences is constructed from the 50 %-identical TEM-1 and Proteus mirabilis beta-lactamases. These hybrids match the TEM-1 sequence except for a region at the C-terminal end, where they are random composites of the two parents. All of these hybrids are biologically inactive. In both experiments, complete loss of activity demonstrates the importance of sequence context in determining whether substitutions are functionally acceptable. Contrary to the prevalent view, then, enzyme function places severe constraints on residue identities at positions showing evolutionary variability, and at exterior non-active-site positions, in particular. Homologues sharing less than about two-thirds sequence identity should probably be viewed as distinct designs with their own sets of optimising features.
J Mol Biol 2000 Aug 18
PMID:Extreme functional sensitivity to conservative amino acid changes on enzyme exteriors. 1096 72

The purpose of the present study was to determine whether Octopus vulgaris spermatozoa are activated by progesterone stimulation. Spermatozoa were collected from the spermatophores in the Needham's sac of the male (MS) and from the spermathecae of oviducal glands of the female (FS). We used transmission (TEM) and scanning (SEM) electron microscopy to study the morphology of untreated, Ca2+ ionophore A23187 and progesterone-treated MS spermatozoa, and untreated FS spermatozoa. We showed that ionophore and progesterone stimulation of MS spermatozoa induce breakdown of the membranes overlapping the acrosomal region, exposing the spiralized acrosome. These modifications resemble the acrosome reaction observed in other species. FS stored in the spermathecae did not show the membranes covering the acrosomal region present in the MS spermatozoa. When ionophore and progesterone treatments were performed in Ca2+-free artificial sea water, no changes were observed, suggesting the role of external calcium in modifying membrane morphology. Lectin studies showed a different fluorescence distribution and membrane arrangement of FS-untreated spermatozoa with respect to the MS, suggesting that spermatozoa transferred in the female genital tract after mating, are stored in a pre-activated state. The plasma membrane of the untreated MS and FS spermatozoa was labelled with Progesterone-BSA-FITC, indicating the presence of plasma membrane progesterone receptor. Taken together these data suggest that progesterone induces an acrosome- like reaction in MS spermatozoa similar to that induced by calcium elevation. In addition progesterone may play a role in the pre-activation of spermatozoa stored in the female tract, further supporting the hypothesized parallelism between cephalopods and vertebrates.
Mol Reprod Dev 2001 May
PMID:Progesterone induces activation in Octopus vulgaris spermatozoa. 1133 51

Degeneration of human male germ cells was analysed by means of light (LM) and transmission electron (TEM) microscopy. The frequency of degenerating cells was correlated with that of Fas-expressing germ cells in human testes with normal spermatogenesis (n = 10), complete early maturation arrest (EMA) (n = 10) or incomplete late maturation arrest (LMA; n = 10) of spermatogenesis. LM analysis of testis sections with normal spermatogenesis indicated that degenerating germ cells were localized in the adluminal compartment of the seminiferous epithelium. TEM showed that apoptotic cells were mostly primary spermatocytes and, to a lesser extent, round or early elongating spermatids. Apoptotic germ cells appeared to be eliminated either in the seminiferous lumen or by Sertoli cell phagocytosis. An increased number of degenerating cells was observed in testes with LMA as compared with normal testes and testes with EMA of spermatogenesis (P < 0.001, Wilcoxon's rank sum test). Comparison of these results with those obtained from immunohistochemistry experiments demonstrated a tight correlation between the number of apoptotic cells and the number of Fas-expressing germ cells (P = 0.001, Spearman's rank = 0.69). These findings suggest that altered meiotic and post-meiotic germ cell maturation might be associated with an up-regulation of Fas gene expression capable of triggering apoptotic elimination of defective germ cells.
Mol Hum Reprod 2002 Mar
PMID:Fas expression correlates with human germ cell degeneration in meiotic and post-meiotic arrest of spermatogenesis. 1187 Feb 28

The structure of a chromosomal extended-spectrum beta-lactamase (ESBL) having the ability to hydrolyze cephalosporins including cefuroxime and ceftazidime has been determined by X-ray crystallography to 1.75 A resolution. The species-specific class A beta-lactamase from Proteus vulgaris K1 was crystallized at pH 6.25 and its structure solved by molecular replacement. Refinement of the model resulted in crystallographic R and R(free) of 16.9 % and 19.3 %, respectively. The folding of the K1 enzyme is broadly similar to that of non-ESBL TEM-type beta-lactamases (2 A rmsd for C(alpha)) and differs by only 0.35 A for all atoms of six conserved residues in the catalytic site. Other residues promoting extended-spectrum activity in K1 include the side-chains of atypical residues Ser237 and Lys276. These side-chains are linked by two water molecules, one of which lies in the position normally filled by the guanidinium group of Arg244, present in most non-ESBL enzymes but absent from K1. The ammonium group of Lys276, ca 3.5 A from the virtual Arg244 guanidinium position, may interact with polar R2 substitutents on the dihydrothiazene ring of cephalosporins.
J Mol Biol 2002 Mar 15
PMID:Structure of an extended-spectrum class A beta-lactamase from Proteus vulgaris K1. 1191 82

Recently, Quintana and others published the results of TEM investigations of ferritin cores extracted from the brain tissue of patients suffering from progressive supranuclear palsy (PSP) and Alzheimer's disease (AD). These ferrihydrite (fFe2O3.9H2O) cores from the iron storage protein ferritin were found to contain a ferrous (Fe2+) iron-bearing biomineral with cubic structure similar to the magnetite standards which were measured. This is a highly important result as magnetic and electron microscopy analysis has demonstrated the presence of magnetite (Fe3O4) and maghemite (gammaFe2O3) in human brain tissue.
Cell Mol Biol (Noisy-le-grand) 2001
PMID:On the structural form of iron in ferritin cores associated with progressive supranuclear palsy and Alzheimer's disease. 1087 42

Pressured by antibiotic use, resistance enzymes have been evolving new activities. Does such evolution have a cost? To investigate this question at the molecular level, clinically isolated mutants of the beta-lactamase TEM-1 were studied. When purified, mutant enzymes had increased activity against cephalosporin antibiotics but lost both thermodynamic stability and kinetic activity against their ancestral targets, penicillins. The X-ray crystallographic structures of three mutant enzymes were determined. These structures suggest that activity gain and stability loss is related to an enlarged active site cavity in the mutant enzymes. In several clinically isolated mutant enzymes, a secondary substitution is observed far from the active site (Met182-->Thr). This substitution had little effect on enzyme activity but restored stability lost by substitutions near the active site. This regained stability conferred an advantage in vivo. This pattern of stability loss and restoration may be common in the evolution of new enzyme activity.
J Mol Biol 2002 Jun 28
PMID:Evolution of an antibiotic resistance enzyme constrained by stability and activity trade-offs. 1207 36

By mixing an aqueous solution of CuCl2 with an NaDC aqueous solution of various concentration and initial molar ratio, seven coordinated samples with distinct appearances and characters were obtained. Their structures and components were investigated by FT-IR spectroscopy, EXAFS (the extended X-ray absorption fine structure), thermal analysis, X-ray diffraction, laser light scattering, TEM (transmission electron micrograph), element analysis and ICP (inductively coupled plasma) analysis. The following conclusions were given: (1) The complexes of Cu2+-NaDC with distinct appearances and properties were synthesized. (2) After Cu(DC)2 dissolved in NaDC aqueous solution, larger micelles (30-90 nm diameter) formed in the supernate, it is a mixed micelle with Cu(DC)2 and NaDC. So these micelles are a new kind of micelle containing two kinds of metal ions. This is a new result using metal ions as bridges to form micelle. (3) According to the different concentration of Cu2+ to NaDC, the complexes formed as gel or poly-crystals. Both the composition of gel complexes and the coordination structure of carboxyl groups with metal ions varied with the initial molar ratio of Cu2+ to Na+. The gel complexes exhibits the non-stoichiometric character. (4) These results are in agreement with physiological condition. All the different states such as gel, precipitate, micelles of various structures are present in bile of gallbladder. We can suggest an ideal model of the interaction between Cu2+ and bile salts in vivo.
Spectrochim Acta A Mol Biomol Spectrosc 2002 May
PMID:The interaction of Cu2 + ions and NaDC micelles. 1208 72

We have previously reported on the IR spectra of styrene, acrylonitrile, and methyl methacrylate (known as the SAN co-polymer), as well as the glass transitions. Not too much work on the refractive indices has been performed, and the refractive indices for a variety of these mixtures and some selected TEM pictures will be presented in this note.
Spectrochim Acta A Mol Biomol Spectrosc 2002 Oct
PMID:Refractive indices for various co-polymer mixtures of styrene, acrylonitrile, and methyl methacrylate. 1239 47

A robust and fast DNA chip method was developed in order to detect the various beta-lactam antibiotic-resistance genes in one slide. These genes included PSE, OXA, FOX, MEN, CMY, TEM, SHV, OXY, and AmpC. beta-lactam antibiotic-resistance genes were labeled with a fluorescent nucleotide by a multiplex polymerase chain reaction using a mixture of specific primer sets for each gene. This labeled target was hybridized with a DNA chip that contained the spots of the specific probe DNAs for each beta-lactam antibiotic-resistance gene. This technique made it possible to detect the specific resistance gene, even in a single bacterium.
Mol Cells 2002 Oct 31
PMID:Development of DNA chip for the simultaneous detection of various beta-lactam antibiotic-resistant genes. 1244 90


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>