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

Somatic extracts of Nippostrongylus brasiliensis contain protease inhibitor(s) capable of inhibiting the activity of trypsin and chymotrypsin A and B. This inhibitor was partially purified by affinity chromatography. Its molecular weight is in the range of 9500-10 000. The inhibition of both trypsin and chymotrypsin depends on the same or closely adjacent active sites of the inhibitor molecule. The inhibitor is unaffected by heating, pH changes or urea, but is sensitive to 2-mercaptoethanol The formation of the enzyme-inhibitor complex is time-dependent. The complex does not dissociate with KC1. The inhibitor has no effect on the activity of elastase, subtilisin, pepsin, rennin, papain and collagenase.
Mol Biochem Parasitol 1981 Jun
PMID:A protease inhibitor of Nippostrongylus brasiliensis. 725 48

It is shown that to obtain the protein-derived basic CD spectra for alpha-helical, beta-structural and irregular regions it is necessary to use a common criterion for isolation of secondary structures for all the reference proteins. Using the "rigid" criterion proposed by Finkelstein, Ptitsyn, Kozytsyn and the "mild" one (as proposed by Levitt and Greer) isolation of the alpha- and beta-structural regions for 5 reference proteins (myoglobin, lysozyme, ribonuclease A, papain, lactate dehydrogenase) has been carried out. Using the f alpha, f beta and f irregular thus obtained and the experimental CD spectra of these proteins, the basic CD spectra for alpha-helical, beta-structural and irregular regions in the proteins have been calculated. It is shown that the use of criterions common for all the proteins leads to good agreement between the calculated basic CD spectra for alpha-helical, beta-structural and irregular forms and the CD spectra of polylysine in the corresponding conformations. The secondary structure of the proteins studied has been analysed using the new protein-derived basic spectra and fairly good quantitative agreement with the X-ray data was achieved.
Mol Biol (Mosk)
PMID:[Determination of the secondary structure of proteins from their circular dichroism spectra. I. Protein reference spectra for alpha-, beta- and irregular structures]. 742 9

Fab fragments from Jel 103, an antibody which specifically binds to single-stranded poly(rl), were prepared by papain digestion, separated into eight isoforms and characterized by mass spectrometry. One of the purified isoforms yielded crystals suitable for structural studies by X-ray diffraction and its crystal structure was determined to 2.4 A resolution. Soaking the crystals in solutions containing either of the mononucleotides inosine-5'-diphosphate, guanosine-5'-diphosphate or deoxyinosine-5'-monophosphate resulted in binding of the nucleotide in a single binding site. However, adenosine-5'-diphosphate does not bind to this antibody. The recognition of the base is achieved through hydrogen bonds to the C6 carbonyl oxygen and the imino NH group of the purine in a pattern similar to that of the base-base interactions in a double-stranded nucleic acid. Additional binding energy is provided by stacking of the base and the Tyr32L side-chain and by interaction of the alpha-phosphate with the antibody in an anionic binding site. Most of the side-chains interacting with the nucleotide come from the light chain. Surprisingly, this antibody shares the VL sequence with another nucleic acid-binding antibody, BV04-1. The latter binds to a single stranded DNA with a high preference for thymine bases. The structures of the unliganded and complexed Jel 103 Fab are compared to those of BV-04-1 Fab and while they show similarity in recognition of the base of the immunodominant nucleotide, their 5' phosphates occupy different positions, suggesting different orientation of the nucleic acid bound to these two antibodies. Differences in the conformations of the L1 loops between the two Fabs have been noted.
J Mol Biol 1994 Oct 21
PMID:Preparation, characterization and crystallization of an antibody Fab fragment that recognizes RNA. Crystal structures of native Fab and three Fab-mononucleotide complexes. 752 84

Trypanosoma cruzi, a protozoan parasite, is the etiologic agent of American trypanosomiasis or Chagas' disease. Chagas' disease afflicts more than 24 million individuals in South and Central America producing a debilitating life-long disease. It is the leading cause of heart failure in many Latin American countries. Currently, there is no satisfactory treatment for this parasitic infection. Cruzain (also known as cruzipain, gp 57/51), the major cysteine protease present in T. cruzi, is critical for the development and survival of the parasite within the host cells, making this enzyme a target for potential trypanocidal drugs. Here we report the X-ray crystal structure of cruzain complexed with the potent inhibitor Z-Phe-Ala-fluoromethyl ketone. The structure was determined at 2.35 A (Rcryst = 0.15) by molecular replacement using a modified papain as the search model. The refined structure is compared to papain. Features which distinguish cruzain from papain are discussed since they may aid in the design of specificity inhibitors. Fluorescence microscopy shows that a biotinylated form of the bound inhibitor does not effectively reach host proteases in their lysosomal compartment, but is selectively taken up by the parasite. The inhibitor greatly reduces parasitemia in a cell culture system, without adverse effects to mammalian cells. This biological selectivity can be exploited, in conjunction with unique active site features revealed by the crystal structure, to develop chemotherapy for Chagas' disease.
J Mol Biol 1995 Mar 24
PMID:The crystal structure of cruzain: a therapeutic target for Chagas' disease. 770 73

The crystal structure of Clostridium thermocellum endoglucanase CelD revealed an extended NH2-terminal segment (involving residues from the putative leader peptide) sticking out from the enzyme core to interact with a symmetry related molecule through an intermolecular salt bridge (Lys38-Asp201). Enzymatic digestion of CelD with various proteases emphasized the flexibility of the NH2-segment in solution. Proteolytic removal of Lys38 or the substitution of bridge-forming residues by site-directed mutagenesis promoted crystal packing arrangements that differ from that of wild type CelD. Crystals of wild-type CelD (a = 99.3 A c = 191.8 A) are trigonal, space group P3(1)21, with one molecule in the asymmetric unit (form A), whereas crystals of papain-treated CelD (a = 100.4 A, c = 248.7 A), of CelDK38M (a = 100.1 A, c = 248.4 A) and of papain-treated CelDD201A (a = 99.9 A, c = 250.0 A) are trigonal, space group P3(1)21, with two crystallographically independent molecules (form B), and crystals of chymotrypsin-treated CelD (a = 100.0 A, c = 254.3 A) and of CelDD201A (a = 99.8 A, c = 254.7 A) are hexagonal, space group P6(1)22, with one molecule in the asymmetric unit (form C). Only chymotrypsin-treated CelD (which preserves both Lys38 and Asp201) can grow in crystal form A upon macroseeding, indicating that formation of the intermolecular salt bridge is critical for stability of this crystal form. Flexible NH2- and COOH-terminal peptide extensions were found to influence crystal nucleation, but not crystal growth. The crystal structures of papain-treated CelD and chymotrypsin-treated CelD, determined at 3.5 A resolution by molecular replacement techniques, demonstrate that a small change in molecular orientation promoted by Lys38 account for the differences between crystal forms B and C.
J Mol Biol 1995 Apr 28
PMID:Multiple crystal forms of endoglucanase CelD: signal peptide residues modulate lattice formation. 773 36

The influence of myosin regulatory light chains (LC2s) lacking the 2kD N-terminal portions of these chains, on internal organization of myosin heads and on actin-myosin interaction was studied with limited proteolysis and polarized fluorescence methods. For these studies heavy meromyosin (HMM) preparations were used: HMM containing intact LC2s (phosphorylated or dephosphorylated) and HMM containing LC2s lacking the 2kD N-terminal portions (including serine which can be phosphorylated). It was found that the susceptibility of the heavy chain cleavage site to trypsin and the alkali light chain (LC1) site to papain of myosin containing shortened regulatory LC2s, is not dependent on saturation of LC2s with Ca2+ or Mg2+ ions. This is in contrast to the myosin containing intact LC2s where Ca2+ or Mg2+ ion saturation does demonstrate a dependence. Similarly, in spectroscopic experiments, dephosphorylated HMM containing intact LC2s causes decrease or increase of actin filament flexibility depending on whether Mg2+ or Ca2+ are bound to LC2s. Correspondingly, HMM with shortened LC2s induces only increase of actin flexibility despite cations being bound. We conclude that the N-terminal fragment of LC2 is important for ensuring a proper Ca2+ dependent conformation of myosin head in the course of its actin-activated ATP hydrolysis.
Biochem Mol Biol Int 1995 Mar
PMID:Significance of the N-terminal fragment of myosin regulatory light chain for myosin-actin interaction. 777 3

4,4'-Diisothiocyanodihydrostilbene-2,2'-disulphonate (H2DIDS), a bifunctional inhibitor of anion exchange in erythrocytes, reacts with Lys-539 in band 3 at neutral pH and crosslinks to Lys-851 at alkaline pH. The accessibility of H2DIDS-labelled band 3 was determined using an anti-H2DIDS antibody and proteolysis. Competitive enzyme-linked immunosorbent assays (ELISAs) showed that a polyclonal antibody raised against H2DIDS-labelled keyhole limpet hemocyanin bound a variety of stilbene disulphonates in the following order of affinities, H2DIDS having the highest affinity: H2DIDS > 4,4'-diisothiocyanostilbene-2,2'-disulphonate (DIDS) > 4-acetamido-4'-isothiocyanostilbene-2,2'disulphonate (SITS) > 4,4'-dinitrostilbene-2,2'-disulphonate (DNDS) > 4,4'-diaminostilbene-2,2'-disulphonate (DADS). The antibody readily detected mono- or bifunctionally H2DIDS-labelled band 3 and proteolytic fragments on immunoblots. H2DIDS attached to Lys-539 is retained in a 7.5 kDa membrane-associated peptide after papain treatment of ghost membranes while the sequence around Lys-851 is more accessible. The band 3 proteolytic fragments protected by the membrane from proteolysis remained associated as a specific complex with a Stokes radius slightly smaller than the dimeric membrane domain after solubilization in detergent solution and retained 82% of the amino acid content of the membrane domain. Circular dichroism (CD) measurements of this H2DIDS-labelled complex showed that it had a very high helical content (86%). The loops connecting the transmembrane segments in H2DIDS-labelled band 3 are therefore not required to maintain transmembrane helix-helix interactions. Denatured band 3 prelabelled with H2DIDS was more readily immunoprecipitated with the anti-H2DIDS antibody than was native band 3 in detergent solution. Deglycosylation of band 3 or proteolytic cleavage of the extramembranous loops did not enhance immunoprecipitation of H2DIDS-labelled band 3. The stilbene disulphonate inhibitor site is therefore relatively inaccessible and is bound by a bundle of helices in the native band 3 protein.
Mol Membr Biol
PMID:Transmembrane helix-helix interactions and accessibility of H2DIDS on labelled band 3, the erythrocyte anion exchange protein. 779 8

cDNA clones encoding cysteine proteinases from cotyledons of germinated seeds of Vicia sativa L. have been obtained by means of PCR. Degenerate oligonucleotide primers were designed according to conserved amino acid regions of known cysteine proteinases. The deduced amino acid sequences of the cDNA clones encoding VSCYSPR1 and VSCYSPR2 display strong homology to cysteine proteinases of the so called papain superfamily. Northern analyses revealed developmentally regulated expression of both the mRNAs in germinating seeds. The transcripts were shown to be products of two distinct single genes, each exhibiting structural polymorphisms as exposed in few nucleotide substitutions.
Plant Mol Biol 1994 Nov
PMID:PCR cloning and expression analysis of cDNAs encoding cysteine proteinases from germinating seeds of Vicia sativa L. 781 78

An alignment/phylogeny of the papain superfamily of cysteine proteases was created using an initial structure-based alignment followed by successive iterations of sequence alignment and phylogenetic inference. The iterative approach resulted in significant improvements in the alignment/phylogeny. There were three groups of cysteine proteases that were distantly related and which could be aligned against each other only in the active site regions: the papain group, which included such stereotypical cysteine proteases as cathepsins B, C, H, L and S; and the bleomycin hydrolase and calpain groups. There was one bacterial sequence in each of the bleomycin hydrolase and calpain groups. The former probably arose by lateral gene transfer, the latter possibly by direct evolution from an ancestral protease predating the eukaryote/prokaryote divergence. The phylogeny of the papain group indicated that many families diverged almost simultaneously early during eukaryotic evolution. In mammals there are at least 12 distinct families of cysteine proteases, possibly many more, including at least two as yet uncharacterized enzymes.
J Mol Biol 1995 Feb 17
PMID:Alignment/phylogeny of the papain superfamily of cysteine proteases. 786 79

The three-dimensional solution structure of recombinant human stefin A has been determined by a simulated annealing protocol using a total of 1113 distance and angle constraints obtained from 1H and 15N HMR spectroscopy. The solution structure is represented by a family of 17 conformers with an average root-mean-square deviation relative to the mean structure of 0.44 A for backbone atoms and 0.94 A for all heavy atoms for the main body of the structure. The protein has a well-defined global fold consisting of five anti-parallel beta-strands wrapped around a central five-turn alpha-helix. There is considerable similarity between the structural features of free stefin A in solution and the X-ray structure of the homologous protein stefin B in its complex with papain, but there are also some important differences in the regions which are fundamental to proteinase binding. The differences consist primarily of two regions of high conformational heterogeneity in free stefin A which correspond in stefin B to two of the components of the tripartite wedge that docks into the active site of the target proteinase. These regions, which are shown to be mobile in solution, are the five N-terminal residues and the second binding loop. In the bound conformation of stefin B they form a turn and a short helix, respectively.
J Mol Biol 1995 Feb 17
PMID:The three-dimensional solution structure of human stefin A. 786 84


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