Starch-Processing Enzymes Produced by Recombinant Bacteria

Exposure of plasmid recombinant microorganisms to an open environment, either inadvertently or intentionally, requires research into those fundamental processes that govern plasmid retention, transfer and expression. In the open environment, a majority of the microbial activity occurs associated with an interface, within thin biological layers consisting of cells and their insoluble extracellular polymer, layers known as biofilms. Current toxic wastewater or wastegas treatment reactors exploit bacterial biofilm systems for certain system operating advantages. Using recombinant bacteria within a biofilm reactor to degrade xenobiotic wastes requires finding a suitable host to harbor and express the desired plasmid phenotype. Suitable host characteristics include: the ability to produce copious amounts of biofilm, resistance to waste-related injury and toxicity, and the ability to retain and express the desired plasmid during long term operation. This paper reports on a laboratory evaluation of factors governing plasmid retention and the expression of trichloroethene (TCE) degradative capacity in both suspended and biofilm cultures.

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Characterization and Site-specific Mutagenesis of the Calcium-binding Protein Oncomodulin Produced by Recombinant Bacteria

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)94090-1 ◽

1989 ◽

Vol 264 (6) ◽

pp. 3470-3477

Author(s):

J P MacManus ◽

C M L Hutnik ◽

B D Sykes ◽

A G Szabo ◽

T C Williams ◽

...

Keyword(s):

Calcium Binding ◽

Binding Protein ◽

Calcium Binding Protein ◽

Recombinant Bacteria ◽

Site Specific

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Repair of DNA Strand Breaks by the Overlapping Functions of Lesion-Specific and Non-Lesion-Specific DNA 3′ Phosphatases

Molecular and Cellular Biology ◽

10.1128/mcb.21.21.7191-7198.2001 ◽

2001 ◽

Vol 21 (21) ◽

pp. 7191-7198 ◽

Cited By ~ 54

Author(s):

John R. Vance ◽

Thomas E. Wilson

Keyword(s):

Dna Damage ◽

Oxidative Dna Damage ◽

Synthetic Lethality ◽

Dna Strand Breaks ◽

Triple Mutant ◽

Phosphatase Domain ◽

Strand Breaks ◽

Alternative Pathways ◽

Processing Enzymes ◽

Dna Strand

ABSTRACT In Saccharomyces cerevisiae, the apurinic/apyrimidinic (AP) endonucleases Apn1 and Apn2 act as alternative pathways for the removal of various 3′-terminal blocking lesions from DNA strand breaks and in the repair of abasic sites, which both result from oxidative DNA damage. Here we demonstrate that Tpp1, a homologue of the 3′ phosphatase domain of polynucleotide kinase, is a third member of this group of redundant 3′ processing enzymes. Unlike Apn1 and Apn2, Tpp1 is specific for the removal of 3′ phosphates at strand breaks and does not possess more general 3′ phosphodiesterase, exonuclease, or AP endonuclease activities. Deletion ofTPP1 in an apn1 apn2 mutant background dramatically increased the sensitivity of the double mutant to DNA damage caused by H2O2 and bleomycin but not to damage caused by methyl methanesulfonate. The triple mutant was also deficient in the repair of 3′ phosphate lesions left by Tdp1-mediated cleavage of camptothecin-stabilized Top1-DNA covalent complexes. Finally, the tpp1 apn1 apn2 triple mutation displayed synthetic lethality in combination with rad52, possibly implicating postreplication repair in the removal of unrepaired 3′-terminal lesions resulting from endogenous damage. Taken together, these results demonstrate a clear role for the lesion-specific enzyme, Tpp1, in the repair of a subset of DNA strand breaks.

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3073. Methylxanthines and the hepatic processing enzymes

Food and Cosmetics Toxicology ◽

10.1016/s0015-6264(76)80240-4 ◽

1976 ◽

Vol 14 (5) ◽

pp. 513-514

Keyword(s):

Processing Enzymes

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Activation of the Kexin from Schizosaccharomyces pombeRequires Internal Cleavage of Its Initially Cleaved Prosequence

Molecular and Cellular Biology ◽

10.1128/mcb.18.1.400 ◽

1998 ◽

Vol 18 (1) ◽

pp. 400-408 ◽

Cited By ~ 21

Author(s):

Dale Powner ◽

John Davey

Keyword(s):

Cleavage Site ◽

Secretory Pathway ◽

Catalytic Domain ◽

Activation Process ◽

Processing Enzymes ◽

Free Translation ◽

A Cell ◽

Internal Site ◽

Primary Cleavage ◽

Full Activation

ABSTRACT Members of the kexin family of processing enzymes are responsible for the cleavage of many proproteins during their transport through the secretory pathway. The enzymes themselves are made as inactive precursors, and we investigated the activation process by studying the maturation of Krp1, a kexin from the fission yeastSchizosaccharomyces pombe. Using a cell-free translation-translocation system prepared from Xenopuseggs, we found that Krp1 is made as a preproprotein that loses the presequence during translocation into the endoplasmic reticulum. The prosequence is also rapidly cleaved in a reaction that is autocatalytic and probably intramolecular and is inhibited by disruption of the P domain. Prosequence cleavage normally occurs at Arg-Tyr-Lys-Arg102↓ (primary cleavage site) but can occur at Lys-Arg82 (internal cleavage site) and/or Trp-Arg99 when the basic residues are removed from the primary site. Cleavage of the prosequence is necessary but not sufficient for activation, and Krp1 is initially unable to process substrates presented in trans. Full activation is achieved after further incubation in the extract and is coincident with the addition of O-linked sugars. O glycosylation is not, however, essential for activity, and the crucial event appears to be cleavage of the initially cleaved prosequence at the internal site. Our results are consistent with a model in which the cleaved prosequence remains noncovalently associated with the catalytic domain and acts as an autoinhibitor of the enzyme. Inhibition is then relieved by a second (internal) cleavage of the inhibitory prosequence. Further support for this model is provided by our finding that overexpression of a Krp1 prosequence lacking a cleavable internal site dramatically reduced the growth rate of otherwise wild-type S. pombecells, an effect that was not seen after overexpression of the normal, internally cleavable, prosequence or prosequences that lack the Lys-Arg102 residues.

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Proteolytic Events of Wound-Healing — Coordinated Interactions Among Matrix Metalloproteinases (MMPs), Integrins, and Extracellular Matrix Molecules

Critical Reviews in Oral Biology & Medicine ◽

10.1177/10454411010120050201 ◽

2001 ◽

Vol 12 (5) ◽

pp. 373-398 ◽

Cited By ~ 144

Author(s):

Bjorn Steffensen ◽

Lari Häkkinen ◽

Hannu Larjava

Keyword(s):

Extracellular Matrix ◽

Wound Healing ◽

Matrix Metalloproteinases ◽

Wound Repair ◽

Enzyme Activation ◽

Processing Enzymes ◽

The Matrix ◽

Signaling Receptors ◽

State Of Rest ◽

And Function

During wound-healing, cells are required to migrate rapidly into the wound site via a proteolytically generated pathway in the provisional matrix, to produce new extracellular matrix, and, subsequently, to remodel the newly formed tissue matrix during the maturation phase. Two classes of molecules cooperate closely to achieve this goal, namely, the matrix adhesion and signaling receptors, the integrins, and matrix-degrading and -processing enzymes, the matrix metalloproteinases (MMPs). There is now substantial experimental evidence that blocking key molecules of either group will prevent or seriously delay wound-healing. It has been known for some time now that cell adhesion by means of the integrins regulates the expression of MMPs. In addition, certain MMPs can bind to integrins or other receptors on the cell surface involved in enzyme activation, thereby providing a mechanism for localized matrix degradation. By proteolytically modifying the existing matrix molecules, the MMPs can then induce changes in cell behavior and function from a state of rest to migration. During wound repair, the expression of integrins and MMPs is simultaneously up-regulated. This review will focus on those aspects of the extensive knowledge of fibroblast and keratinocyte MMPs and integrins in biological processes that relate to wound-healing.

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