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I Palva

Publications and source records attributed to I Palva.

49 records · Page 3Linked to original sources
PubMed1981

Structural complexity of defective-interfering RNAs of Semliki Forest virus as revealed by analysis of complementary DNA.

The 18S defective interfering RNA of Semliki Forest virus has been reverse transcribed to cDNA, which was shown to be heterogeneous by restriction enzyme analysis. After transformation to E.coli, using pBR322 as a vector, two clones, pKTH301 and pKTH309 with inserts of 1.7 kb and 2 kb, were characterized, respectively. The restriction maps of the two clones were different but suggested that both contained repeating units. At the 3' terminus, pKTH301 had preserved 106 nucleotides and pKTH309 102 nucleotides from the 3' end of the viral 42S genome. The conserved 3' terminal sequence was joined to a different sequence in the two clones, and these sequences were not derived from the region coding for the viral structural proteins. The DI RNAs represented by the two clones are generated from the viral 42S RNA by several noncontinuous internal deletions, since the largest colinear regions with 42S RNA are 320 nucleotides in pKTH301, and 430 and 340 nucleotides in pKTH309. All these fragments had unique RNase T1 oligonucleotide fingerprints, suggesting that they were derived from different regions of 42S RNA.

Base Composition↗
PubMed1981

Nucleotide sequence of the promoter and NH2-terminal signal peptide region of the alpha-amylase gene from Bacillus amyloliquefaciens.

We have isolated and partially sequenced the gene coding for alpha-amylase (EC 3.2.1.1) from Bacillus amyloliquefaciens by molecular cloning in the plasmid pUB110 using Bacillus subtilis as a host. The nucleotide sequence of the NH2-terminal region of the cloned gene was determined and found to contain a 31-residue-long stretch of amino acids preceding the NH2-terminal sequence of the extracellular alpha-amylase. Within this sequence there is a 15-residue-long stretch of uncharged amino acids similar to that found at the NH2 terminus of other precursors to exported proteins. This "signal sequence" is probably removed in conjunction with the translocation of alpha-amylase through the cytoplasmic membrane. In vitro labeling of alpha-amylase with radioactive amino acids in a coupled transcription-translation system followed by partial sequencing established the exact location of the NH2 terminus of the alpha-amylase gene. The nucleotide sequence preceding the NH2 terminus has properties resembling the RNA-polymerase- and ribosome-binding sites found at the 5' terminus of many prokaryotic genes.

Amylases↗
PubMedDate not supplied

Molecular cloning of alpha-amylase gene from Bacillus amyloliquefaciens and its expression in B. subtilis.

The gene coding for alpha-amylase from Bacillus amyloliquefaciens was isolated by direct shotgun cloning using B. subtilis as a host. The genome of B. amyloliquefaciens was partially digested with the restriction endonuclease MboI and 2- to 5-kb fragments were isolated and joined to plasmid pUB110. Competent B. subtilis amylase-negative cells were transformed with the hybrid plasmids and kanamycin-resistant transformants were screened for the production of alpha-amylase. One of the transformants producing high amounts of alpha-amylase was characterized further. The alpha-amylase gene was shown to be present in a 2.3-kb insert. The alpha-amylase production of the transformed B. subtilis could be prevented by inserting lambda DNA fragments into unique sites of EcoRI, HindIII and KpnI in the insert. Foreign DNA inserted into a unique ClaI site failed to affect the alpha-amylase production. The amount of alpha-amylase activity produced by this transformed B. subtilis was about 2500-fold higher than that for the wild-type B. subtilis Marburg strain, and about 5 times higher than the activity produced by the donor B. amyloliquefaciens strain. Virtually all of the alpha-amylase was secreted into the culture medium. The secreted alpha-amylase was shown to be indistinguishable from that of B. amyloliquefaciens as based on immunological and biochemical criteria.

Amylases↗
PubMedDate not supplied

Secretion of interferon by Bacillus subtilis.

Bacillus subtilis was transformed with a hybrid gene in which the sequence encoding the alpha-amylase signal peptide was joined by a linker to the sequence encoding mature human interferon alpha 2(IFN-alpha 2). The hybrid preprotein was cleaved precisely following the last amino acid of the alpha-amylase signal sequence and was secreted at 0.5--1 mg per liter. IFN-alpha 2, preceded by either one or six amino acids, has the same specific antiviral activity as IFN-alpha 2 itself.

Amino Acid Sequence↗
PubMedDate not supplied

The secretory S complex in Bacillus subtilis is identified as pyruvate dehydrogenase.

We have cloned the operon for the Bacillus subtilis S complex, which has been suggested to be a component of the protein secretion machinery. The S-complex operon was found to encode 4 proteins, which were identified as subunits of pyruvate dehydrogenase (PDH). The Staphylococcus aureus membrane-bound ribosome protein (MBRP) complex has been considered to be a counterpart of the B. subtilis S complex. Here, we sequenced a fragment of the MBRP operon encoding the C-terminal part of E1 beta, the entire E2 and the N-terminal part of the E3 subunit of PDH, thus conclusively confirming the PDH identity of the MBRP complex as well. It appeared unlikely that PDH could be a primary component in protein secretion, thus disproving the previous hypothesis of the role of the S complex. However, attachment of the S complex (PDH) to the membrane and ribosomes may produce a biologically significant interaction.

Bacillus subtilis↗