Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.31.1 (micrococcal nuclease)
 2,818 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Alu-like elements comprise the most abundant family of interspersed repetitive sequences in primates and rodents, and contain many features of processed genes, suggesting that they were initially derived by reverse transcription of processed RNA transcripts. Transcripts containing Alu family members are represented in heterologous nuclear RNAs, cytoplasmic messenger RNAs and small RNAs, although nothing is known about their function. Evolutionary studies strongly suggest that the parent RNA for the Alu-like elements is the highly conserved 7SL RNA, which is an essential component of signal recognition particle (SRP), a small cytoplasmic ribonucleoprotein whose function is the targeting of nascent secretory and membrane proteins to the rough endoplasmic reticulum (for a review see ref. 6). 7SL RNA is composed of both unique and Alu-like sequences. SRP is rod-shaped and, in addition to its RNA, contains four proteins (two monomers composed of a polypeptide of relative molecular mass (Mr) 19,000 (19K) and one of 54K, and two heterodimers, one composed of a 9K and a 14K polypeptide, and the other composed of a 68K and a 72K polypeptide, respectively). The RNA moiety is required for SRP activity, as well as for structural integrity of the particle. To investigate whether the Alu-like segments of 7SL RNA have a specific role in SRP activity, we have now purified and analysed a SRP subparticle that is created upon extensive digestion with micrococcal nuclease and entirely lacks the Alu-like sequences. We find that it contains the 72/68K, 54K and 19K proteins tightly bound, but lacks the 9/14K protein. In vitro activity assays demonstrated that the subparticle could still promote secretory protein translocation across the microsomal membrane, but could no longer trigger an arrest of pre-secretory protein synthesis. Re-addition of the 9/14K protein did not restore the elongation arrest. We conclude that the region of SRP comprised of the Alu-like RNA and the 9/14K protein exists in a distinct structural domain which is not required for the protein translocation promoted by SRP but apparently confers elongation-arresting activity on the particle.
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PMID:Removal of the Alu structural domain from signal recognition particle leaves its protein translocation activity intact. 241 65

The biological activities of pancreatic presecretory and secretory proteins synthesized in vitro were compared in studies of (a) the binding of nascent amylase to its substrate, glycogen, (b) the binding of nascent trypsinogen 1, trypsinogen 2+3, and chymotrypsinogen 1 to Sepharose-bound soybean trypsin inhibitor, and (c) the activation of nascent trypsinogen by porcine enterokinase. Nascent secretory proteins synthesized in vitro using a mRNA-dependent gel-filtered reticulocyte lysate translation system supplemented with canine pancreas rough microsomes or canine pancreas mRNA and micrococcal nuclease-treated microsomal membranes showed biological activities similar to authentic secretory proteins if oxidized glutathione was added during their synthesis. Proteins synthesized in the presence of membranes and the absence of glutathione showed significantly less biological activity due to incorrect development of conformation. Presecretory proteins synthesized in vitro with canine pancreas mRNA in the absence of microsomal membranes had little or no activity after translation in either the absence or presence of glutathione. These and previous findings (Scheele, G. A., and Jacoby, R. (1982) J. Biol. Chem. 257, 12277-12282) indicate that proteolytic removal of the NH2-terminal transport peptide is necessary to allow correct conformational development, including the formation of native disulfide bonds, which not only stabilizes the molecule but allows expression of authentic biological and probiological activity.
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PMID:Proteolytic processing of presecretory proteins is required for development of biological activities in pancreatic exocrine proteins. 633 49

Isolated rat liver cell nuclei were incubated with the alkylating cytostatic drug cyclophosphamide (CPA) in the presence of a microsomal activation system. Digestion of the 3H-CPA-treated nuclei with DNase I and micrococcal nuclease, respectively, showed that the CPA-modified DNA apparently has become resistant against such enzymatic attack. For analysis of the DNA-CPA reaction products, the DNA was isolated under mild conditions and degraded enzymatically. In cell nuclei whose DNA had been prelabeled with 32P in vivo, a predominant binding of 3H-CPA (about 50%) to the DNA phosphate groups was observed. Terminal phosphate groups apparently play an important role in this reaction. Neutral heating of DNA from 3H-CPA-treated nuclei liberated up to 60% of the initially bound 3H-radioactivity. The results are discussed in relation to the recent data concerning the route of decomposition of activated CPA to phosphoramide mustard and acrolein.
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PMID:Interaction of cyclophosphamide with DNA in isolated rat liver cell nuclei. 671 77

After treatment at a microsomal nuclease concentration too low to reduce the endogenous amino acid-incorporating activity of freshly prepared reticulocyte lysate, there is little, if any, intact 26 S RNA left in the ribosomes of either wheat germ or rabbit reticulocyte cell-free protein synthesizing extracts. The primary scissions, probably at highly exposed sites in the rRNA of plant and animal ribosomes, produce two fragments which remain complexed until thermal denaturation reveals "hidden breaks." Molecular weights of the fragments are approximately 0.5 x 10(6) and 0.8 x 10(6) in the case of wheat, and 0.4 x 10(6) and 1.3 x 10(6) in the case of rabbit. There is little perceptible degradation of 5 S, 5.8 S, and 18 S rRNA, or of tRNA in the same extracts. Even though limited degradation of 26 S rRNA by a reticulocyte nuclease has been reported to severely impair the translational mechanism in reticulocyte ribosomes, micrococcal nuclease-induced degradation of rRNA, whether limited or extensive, does not seriously impair the ability of reticulocyte lysates to discriminate, by selective polypeptide synthesis, between complex populations of cellular mRNA. In an allied study, it is shown that under conditions well suited to recovery of the 5.8 S/26 S rRNA complex, with its naturally occurring hidden break, 5 S/18 S rRNA complexing is not detectable in the RNA or metabolizing embryos, nor in the RNA from untreated or nuclease-treated protein synthesizing extracts from wheat and rabbit. The significance of this finding is briefly elaborated in relation to the suggestion that 5 S rRNA may interact with the M2(6)A-m2(6)A hairpin near the 3'-end of 18 S rRNA.
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PMID:Structural integrity of DNA and translational integrity of ribosomes in nuclease-treated cell-free protein synthesizing systems prepared from wheat germ and rabbit reticulocytes. 724 Jan 74

The mechanism by which secretory proteins are segregated within the cisternal space of microsomal vesicles was studied using dog pancreas mRNA which directs the synthesis of 14 well-characterized nonglycosylated pancreatic exocrine proteins. In the absence of microsomal membranes, each of the proteins was synthesized as larger polypeptide chains (presecretory proteins). 1,000-2,000 daltons larger than their authentic counterparts as judged by polyacrylamide gel electrophoresis in SDS. Conditions optimal for the study of reconstituted rough microsomes in the reticulocyte lysate system were examined in detail using mRNA and microsomal membranes isolated from dog pancreas. Functional reconstitution of rough microsomes was considerably more efficient in the presence of micrococcal nuclease- treated membranes than in the presence of EDTA-treated membranes. Analysis for segregation of nascent secretory proteins by microsomal vesicles, using post-translational incubation in the presence of trypsin and chymotrypsin, 50 mug/ml each, was shown to be inadequate, because of the disruption of vesicles by protease activity. Addition of 1-3 mM tetracaine or 1 mM dibucaine stabilized microsomal membranes incubated in the presence of trypsin and chymotrypsin at either 0 degrees or 22 degrees C. Each of the pancreatic presecretory proteins studied was correctly processed to authentic secretory proteins by nuclease-treated microsomal membranes, as judged by both one-dimensional and two-dimensional gel electophoresis. Post-translational addition of membranes did not result in either segregation or processing of nascent polypeptide chains. Post- translational proteolysis, carried out in the presence of 3 mM tetracaine, indicated that each of the 14 characterized dog pancreas secretory proteins was quantitatively segregated by nuclease-treated microsomal vesicles. Segregation of nascent secretory proteins was irreversible, since radioactive amylase, as well as the other labeled secretory proteins, remained quantitatively sequestered in microsomal vesicles during a 90-min incubation at 22 degrees C after the cessation of protein synthesis. Studies employing synchronized protein synthesis and delayed addition of membranes indicated that all pancreatic presecretory proteins contain amino terminal peptide extensions. These peptide extensions are shown to mediate the cotranslational binding of presecretory proteins to microsomal membranes and the transport of nascent secretory proteins to the vesicular space. The maximum chain lengths which, during synthesis, allow segregation of nascent polypeptide chains varied between 61 (pretrypsinogen 2 + 3) and 88 (preprocarboxypeptidase A1) amino acid residues among dog pancreas presecretory proteins. Reconstitution studies using homologous and heterologous mixtures of mRNA (dog, guinea pig, and rat pancreas; rat liver) and micrococcal nuclease-treated microsomal membranes (dog, guinea pig, and rat liver; dog pancreas), in the presence of placental ribonuclease inhibitor, suggest that the translocation mechanism described is common to the rough endoplasmic reticulum of all mammalian tissues.
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PMID:Mechanism of compartmentation of secretory proteins: transport of exocrine pancreatic proteins across the microsomal membrane. 746 18