Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.12.7.2 (hydrogenase)
 3,522 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cellular location of membrane-bound NiFe-hydrogenase 2 (HYD2) from Escherichia coli was studied by immunoblot analysis and by activity staining. Treatment of spheroplasts with trypsin was able to release active HYD2 into the soluble fraction, indicating that HYD2 is attached to the periplasmic side of the cytoplasmic membrane and that HYD2 undergoes a trans-membrane translocation during its biosynthesis. By using a nik mutant deficient in the high affinity specific nickel transport system, we show that the intracellular availability of nickel is essential for the processing of the large subunit and for the transmembrane translocation of HYD2. We also demonstrate that the processing of the precursor, which is related with nickel incorporation, can occur in the membrane-depleted soluble fraction and that it is associated with the increase in resistance to proteolysis of the processed form of the large subunit. The mechanism of the transmembrane translocation of HYD2 is discussed.
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PMID:Requirement for nickel of the transmembrane translocation of NiFe-hydrogenase 2 in Escherichia coli. 877 79

Maturation of the large subunit of E. coli hydrogenase 3, HycE, requires the action of seven accessory proteins. The HycI protease catalyses a C-terminal proteolytic cleavage of the large subunit, which was shown to result in a dramatic change in migration behavior of HycE in nondenaturing PAGE. HypA, HypB, HypC, HypD, HypE, and HypF are required for metallocenter assembly. A polyacrylamide gel system under nondenaturing conditions was used for the investigation of any protein-protein interactions between HycE and the Hyp proteins. It revealed the existence of a complex between the precursor of HycE (pre-HycE) with one of the accessory proteins, namely HypC. HypC migrates in at least three different forms in nondenaturing PAGE, the appearance of one of which (form 1) is strictly dependent on the presence of unprocessed HycE in the extract. Overexpression of either hypC or hycE from a plasmid leads to an increased formation of this HypC-form 1. In two-dimensional polyacrylamide gel electrophoresis with nondenaturing PAGE as the first and SDS-PAGE as the second dimension, this HypC form comigrates with part of the pre-HycE protein. This comigration was also observed in anion exchange chromatography. To analyze the pre-HycE-HypC complex in more detail, HypC was overproduced and purified. The purified protein was able to bind to pre-HycE in vitro. These results and also the finding that the processed form of HycE is not associated with HypC suggest that HypC binds to pre-HycE to keep it in a conformation accessible for metal incorporation.
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PMID:Interaction of the hydrogenase accessory protein HypC with HycE, the large subunit of Escherichia coli hydrogenase 3 during enzyme maturation. 948 46