Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0272170 (SDS)
 50,377 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The aminopeptidase activity of a homogenate of rabbit kidney treated with Triton X-100 was measured using L-aminoacyl-2-naphthylamides (AA-NA). After gradient elution ion-exchange chromatography, four peaks of aminopeptidase activity were eluted. The enzyme eluted at 450 microS containing 33.5% of the activity towards Arg-NA was applied to a Superdex 75 column and presented only one protein band on 10% SDS-polyacrylamide gel electrophoresis. This enzyme has an apparent molecular mass of 78 kDa, is five-fold activated by 0.15 M NaCl and the highest Vmax/KM ratio was obtained with Arg-NA. Enzyme activity was inhibited 100% by 0.13 mM sodium p-hydroxymercuribenzoate, 20% by 0.75 mM EDTA and 100% by 0.66 mM o-phenanthroline. Puromycin and bestatin behaved like competitive inhibitors with a Ki of 0.60 mM and 5.0 microM and 5.0 microM, respectively.
 ...
PMID:Basic aminopeptidase from rabbit kidney: purification and partial characterization. 919 41

A protamine-degrading marine bacterium was isolated from marine soil and identified as Aeromonas salmonicida subsp. based on its taxonomical characteristics. An alanine-specific aminopeptidase, called aminopeptidase K, from an extract of the strain was purified and characterized. The aminopeptidase K was purified about 80-fold by fractionation with ammonium sulfate and column chromatography on QA-52 cellulose, Phenyl Superose and Superose 12. The purified enzyme is composed of 6 subunits of 86 kDa with a molecular mass of 520 kDa according to gel filtration and SDS-PAGE. The N-terminal sequence of the enzyme was H.Gly-Gln-Gln-Pro-Gln-Ile-Lys-Try-Tyr-His-Asp-Tyr-Asp-Ala-Pro-Asp-Tyr-Ty r- Ile-Thr-. It is inhibited by monoiodoacetate, N-ethylmaleimide, and puromycin. The Michaelis constant (K(m)) and the maximal rate of hydrolysis (Vmax) were, respectively, 0.28 mM and 49.4 mumol/min/mg for the L-Ala-beta-naphthylamide substrate. The optimum pH and optimum temperature were 6.5 and 45 degrees C, respectively. The purified enzyme was highly specific to L-Ala-beta-naphthylamide.
 ...
PMID:Purification and properties of an aminopeptidase from a protamine-degrading marine bacterium. 925 72

Alkaline extracellular protease (AEP) from Yarrowia lipolytica is synthesized as a precursor with a 157 aa prepro-region. Signal peptide cleavage was shown to occur after Ala15 by N-terminal amino acid radiosequencing of the largest intracellular AEP precursor. AEP proteolytic activity was not required for AEP processing. After a change of the putative active site Ser to Ala, inactive AEP with the same mobility on SDS-PAGE as wild-type mature AEP was secreted. The role of dipeptidyl aminopeptidase (DPAPase) activity in AEP processing was also investigated. Mutations early in the -X-Ala- and -X-Pro- dipeptide stretch (Pro17 to Met which should prevent DPAPase processing and Ala19 to Val which should allow removal of only the first dipeptide) did not prevent synthesis of active mature AEP nor did use of the DPAPase inhibitor ProboroPro. Deletion of the entire dipeptide stretch (Ala16 to Pro33) resulted in intracellular accumulation of an AEP precursor, which surprisingly was not glycosylated, and little or no secretion of AEP-related polypeptides. Expression of AEP in wild-type and dpp1 dap2 Saccharomyces cerevisiae strains (lacking both the Golgi and vacuolar DPAPases) resulted in secretion of only mature AEP and no AEP precursors. Transit times and levels of AEP secretion were similar for both strains. These results indicate that the KEX2-like cleavage after Lys156-Arg157, which yields mature active AEP can occur in the absence of DPAPase processing and that DPAPase processing is not necessary for secretion of mature active AEP.
 ...
PMID:Dipeptidyl aminopeptidase processing and biosynthesis of alkaline extracellular protease from Yarrowia lipolytica. 935 27

The proteasome (EC 3.4.99.46) is a high molecular mass (approximately 700 kDa) multisubunit enzyme complex which is the focus of worldwide research in order to identify the structure, mechanism of action and specificity of the complex. The purpose of the present study was to investigate the tryptic, chymotryptic and peptidylglutamyl-peptide hydrolysing (PGPH) activities of ostrich liver proteasome. The proteasome was purified from ostrich liver by employing ammonium sulphate fractionation, followed by three sequential chromatographic steps on Toyopearl Super Q-650 S, Sephadex G-150 and phenyl-Toyopearl columns. Temperature and pH optima were examined and the effect of inhibitors, detergents, fatty acids and cations on the peptidase activities was determined. Ostrich proteasome exhibited a relative M(r) of approximately 665,000 using non-denaturing gradient PAGE and dissociated into the characteristic "ladder" associated with the proteasome subunits during SDS-PAGE. The pH optima for the peptidase activities were found to be slightly alkaline (tryptic activity) and neutral (chymotryptic-like and PGPH activities). Ostrich liver proteasome was found to be activated in terms of the PGPH activity by fatty acids and SDS, whereas the chymotryptic and tryptic-like activities were differentially inhibited. Ostrich proteasome, in its inhibition by monovalent cations, was similar to the proteasomes extracted from other sources. The specificity of the proteasome appears to be very broad, although it lacks aminopeptidase activity. The yield compared favourably with similar extraction procedures which have been reported. On the basis of the physicochemical and kinetic properties which ostrich liver proteasome exhibited, it can be safely concluded that it corresponds well with the proteasomes isolated from many other sources.
 ...
PMID:Purification and characterization of proteasome from ostrich liver. 936 39

We have identified on the membranes of the locomotory muscle of Ascaris suum an amastatin-sensitive aminopeptidase that hydrolyses the bioactive neuropeptides AF1 (KNEFIRF-NH2) and AF2 (KHEYLRF-NH2), by cleavage of the Lys1-Asn2 and Lys1-His2 peptide bonds, respectively. AF2 (1.2 nmol of HEYLRF-NH2 formed min[-1] (mg protein[-1])) was hydrolysed at a faster rate compared to AF1 (0.2 nmol of NEFIRF-NH2 formed min[-1] (mg protein[-1])). AF1 hydrolysis by the aminopeptidase was inhibited by the amastatin (IC50, 9.0 microM), leuhistin (IC50, 1.25 microM) but was insensitive to puromycin, indicating a similarity to mammalian aminopeptidase N. The enzyme was also inhibited by arphamenine B (IC50, 9.0 microM), (2S, 3R)-3-amino-2-hydroxy-4-(4-nitrophenyl)butanoyl-L-leucine (IC50, 8.0 microM), bestatin (IC50, 15.0 microM) and 1 mM 1-10 bis-phenanthroline. The detergent Triton X-100 solubilised enzyme had a pI of 5.0 and after 1000-fold purification by ion-exchange chromatography, appeared to have a Mr of around 240,000 by SDS-PAGE. The purified aminopeptidase had a Km of 534 microM for the hydrolysis of AF1 and cleaved Phe1 from FMRF-NH2, but was unable to hydrolyse DFMRF-NH2 or FDMRF-NH2. The aminopeptidase that we have described in this report might have a role in the extracellular metabolism and inactivation of neuropeptides acting on the locomotory muscle of A. suum.
 ...
PMID:Purification and properties of a membrane aminopeptidase from Ascaris suum muscle that degrades neuropeptides AF1 and AF2. 936 67

Alanyl aminopeptidase (AAP) was purified to homogeneity from human seminal plasma. The calculated molecular weight of the purified enzyme was approximately 137,000+/-5,000 from light scattering, 140,000 (main) and 137,000 (minor) from non-denatured PAGE and 153,000 from SDS-PAGE in the absence or presence of 2-mercaptoethanol (2-ME). These findings suggest that the enzyme is monomeric in form in human seminal plasma. The enzyme hydrolyzed several amino acid 4-methyl-coumaryl-7-amide (MCA) substrates. The order of Kcat/Km values of AAP at optimal pH (pH 7.5) was Ala- > Lys-Ala- > or = Met- > Leu- > Phe- > Arg- > or = Arg-Arg- > Lys- > Gly-MCAs. AAP was potently inhibited by bestatin, leuhistin, actinonin, amastatin, and 1,10-phenanthroline. These findings suggest that AAP is an aminopeptidase. We determined that the amino acid sequence of the first 22 residues of the enzyme was Ser1-Thr-Thr-Pro-Ser5-Ala-Ser-Ala-Thr-Thr10-Asn-Pro-Al a-Ser-Ala15-Thr-Thr-Leu-Asp-Gln20-Ser-Lys-. This sequence was completely coincident with that downstream of the transmembrane site of human intestinal alanyl aminopeptidase N (CD13). We also isolated cDNA encoding AAP from human prostate cDNA library, sequenced its structure, and confirmed human seminal plasma AAP to be identical with alanyl aminopeptidase N. We postulated that native human seminal plasma alanyl aminopeptidase is released into the seminal plasma after the specific site is cleaved by elastase or an elastase-like enzyme. The enzyme level in human seminal plasma determined by single radial immunodiffusion was 5.2+/-3.2 mg/100 ml (mean+/-SD, n=20) in individuals 20-47 years of age. AAP was immunohistochemically stained in the luminal site-cell membrane of epithelial cells in the prostatic gland and ductuli efferentes of the testis.
 ...
PMID:Alanyl aminopeptidase from human seminal plasma: purification, characterization, and immunohistochemical localization in the male genital tract. 939 82

An intracellular X-prolyl-dipeptidyl aminopeptidase from Streptococcus thermophilus ACA-DC 4, isolated from traditional Greek yoghurt, was purified by anion exchange and gel filtration chromatography. A single band of molecular weight of about 80,000 appeared in SDS-PAGE; by gel filtration it was shown that the native enzyme was dimeric. The peptidase showed optimum activity on glycyl-prolyl 4-nitroanilide at pH 7.0 and at 50 degrees C, with K(m) = 3.1 mM and Vmax = 3500 U mg-1; over 50 degrees C the enzyme activity declined rapidly. It was inactivated by PMSF; sulfhydryl group reagents and metal chelators had little effect on enzyme activity.
 ...
PMID:Purification and characterisation of an intracellular X-prolyl-dipeptidyl aminopeptidase from Streptococcus thermophilus ACA-DC 4. 951 81

Blasticidin S-producing Streptomyces morookaensis JCM4673 produces an enzyme which inactivates puromycin (PM) by hydrolyzing an amide linkage between its aminonucleoside and O-methyl-L-tyrosine moieties [Nishimura et al. (1995) FEMS Microbiol. Lett. 132, 95-100]. In this study, we purified to homogeneity the enzyme from the cell-free extracts of S. morookaensis. The molecular weight of PM-hydrolyzing enzyme, estimated by SDS-PAGE and gel filtration, was 68 and 66 kDa, respectively, suggesting that this protein is monomeric. The PM-hydrolyzing activity was strongly inhibited by Zn2+, Fe2+, Cu2+, Hg2+, and N-bromosuccinimide, but was stimulated by DTT. The optimum pH and temperature for PM-hydrolyzing activity were 8.0 and 45 degrees C, respectively. Several L-aminoacyl-beta-naphthylamides were good substrates for the enzyme, suggesting that the PM-inactivating enzyme has an aminopeptidase activity. The N-terminal sequence of the first 14 amino acids (Val-Ser-Thr-Ala-Pro-Tyr-Gly-Ala-Trp-Gln-Ser-Pro-Ile-Asp) of the enzyme showed no significant homology with any published hydrolase sequences.
 ...
PMID:Purification and characterization of a puromycin-hydrolyzing enzyme from blasticidin S-producing Streptomyces morookaensis. 953 99

An aminopeptidase with a preference for N-terminal aspartyl and glutamyl residues but distinct from glutamyl aminopeptidase (EC 3.4. 11.7) was purified to near homogeneity from rabbit brain cytosol. Its properties were similar to an enzyme described previously (Kelly, J. A., Neidle, E. L., and Neidle, A. (1983) J. Neurochem. 40, 1727-1734). Aspartyl aminopeptidase had barely detectable activity toward simple aminoacyl-naphthylamide substrates. Its activity was determined with the substrate Asp-Ala-Pro-naphthylamide in the presence of excess dipeptidyl-peptidase IV (EC 3.4.14.5). The native enzyme has a molecular mass of 440 kDa and migrates as a single band of 55 kDa after SDS-polyacrylamide gel electrophoresis. The sequences of three tryptic peptides were used to screen the GenBankTM data base of expressed sequence tags. Human and mouse clones described as "similar to a yeast vacuolar aminopeptidase" and containing full-length cDNAs were identified and sequenced. The human cDNA was expressed in Escherichia coli. The amino acid sequence has significant homology to yeast aminopeptidase I, placing it as the first identified mammalian member of the M18 family of metalloproteinases. Homologous sequences in Caenorhabditis elegans and in prokaryotes revealed three conserved histidines, three conserved glutamates and five conserved aspartates. Aspartyl aminopeptidase is found at relatively high levels in all mammalian tissues examined and is likely to play an important role in intracellular protein and peptide metabolism.
 ...
PMID:Purification, characterization, and cloning of a cytosolic aspartyl aminopeptidase. 963 44

Aminopeptidase H was isolated and purified from fresh skeletal muscle of the lizard Agama stellio stellio by ammonium sulfate fractionation and successive chromatographies on DEAE-cellulose, Ultrogel AcA-34, activated thiol-Sepharose 4B, phenyl-Sepharose CL-4B, and DEAE-cellulose again. This is the first report of the isolation of aminopeptidase H from a reptile. The purified enzyme migrated as a single band on SDS-PAGE. The molecular weight of the enzyme was 48 kD by SDS-PAGE and 384 kD on Ultrogel AcA-34 column chromatography. The optimum pH for hydrolysis of L-leucine beta-naphthylamide (Leu-Nap) was 7.8. The Km values for the hydrolysis of Leu-Nap and Nalpha-benzoyl-DL-arginine beta-naphthylamide (BzArg-Nap) were 0.48 and 0.99 mM, respectively. These activities were strongly inhibited by iodoacetic acid and leupeptin but were not affected by EDTA, pepstatin, bestatin, or phenylmethylsulfonyl fluoride. The enzyme has been shown not to hydrolyze proteins such as hemoglobin, BSA, myofibrillar proteins, and sarcoplasmic proteins.
 ...
PMID:Isolation, purification, and properties of aminopeptidase H from skeletal muscle of the lizard Agama stellio stellio. 1018 16


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