Two Bifunctional Enzymes with Ferric Reduction Ability Play Complementary Roles during Magnetosome Synthesis in Magnetospirillum gryphiswaldense MSR-1

Abstract

cThe bacterial strain Magnetospirillum gryphiswaldense MSR-1 does not produce siderophores, but it absorbs a large amount of ferric iron and synthesizes magnetosomes. We demonstrated previously the presence of six types of ferric reductase isozymes (termed FeR1 through FeR6) in MSR-1. Of these isozymes, FeR5 was the most abundant and FeR6 showed the highest ferric reductase activity. In the present study, we cloned the fer5 and fer6 genes from MSR-1 and expressed them separately in Escherichia coli. FeR5 and FeR6 were shown to be bifunctional enzymes through analysis of amino acid sequence homologies, structural predictions (using data from GenBank), and detection of enzyme activities. FeR5 is a thioredoxin reductase and FeR6 is a flavin reductase, in addition to being ferric reductases. To elucidate the functions of the enzymes, we constructed two single-gene-deletion mutant strains (⌬fer5 and ⌬fer6 mutants) and a double-gene-deletion mutant strain (⌬fer5 ⌬fer6 [⌬fer5؉6] mutant) along with its complemented strains (C5 and C6). An evaluation of phenotypic and physiological properties did not reveal significant differences between the wild-type and single-gene-deletion strains, whereas the double-gene-deletion strain showed reduced iron absorption and no magnetosome synthesis. Complementation of the double-gene-deletion strain using either fer5 or fer6 resulted in the partial recovery of magnetosome synthesis. Quantitative real-time PCR analysis of fer5 and fer6 transcriptional levels in the wild-type and complemented strains demonstrated consistent transcription of the two genes and confirmed that FeR5 and FeR6 are bifunctional enzymes that play complementary roles during the process of magnetosome synthesis in MSR-1.

Transcriptome analysis reveals physiological characteristics required for magnetosome formation in Magnetospirillum gryphiswaldense MSR‐1

“…Recombinant non‐magnetosome strain Rhodospirillum rubrum by transferring these operons into successfully synthesized magnetosomes (Kolinko et al ., ). In addition, genes out of MAI also play essential roles in the biomineralization process include fur (encoding ferric uptake regulator Fur) (Uebe et al ., ; Qi et al ., ), feoAB2 (encoding ferrous iron transport protein) (Rong et al ., ) and fer (encoding ferric reductase) (Zhang et al ., ). Denitrification genes also affect magnetosome formation, and particularly nap genes (encoding periplasmic nitrate reductases) involved in redox reactions (Li et al ., ; Raschdorf et al ., ).…”

Expression patterns of key iron and oxygen metabolism genes during magnetosome formation inMagnetospirillum gryphiswaldenseMSR-1

“…At least six ferric reductases are present in MSR‐1 (Meng et al ., ). FeR5 and FeR6 are ferric iron reductases but also display thioredoxin reductase activity and flavin reductase activity, respectively (Zhang et al ., ). We observed increases in the expression of both fer5 (MGR_2378) and fer6 (MGR_3076) during magnetosome maturation, and the expression patterns of these two genes were consistent (Fig.…”

“…c). These findings indicate that FeR5 and FeR6 play complementary roles during the process of magnetosome synthesis in MSR‐1 (Zhang et al ., ) following energy exchange by simultaneous electron transfer and oxidative reactions. In agreement with this concept, Suzuki et al .…”

Natural Attenuation of Mn(II) in Metal Refinery Wastewater: Microbial Community Structure Analysis and Isolation of a New Mn(II)-Oxidizing Bacterium Pseudomonas sp. SK3

“…(all Ms. gryphiswaldense; Table S2) accounted for 3.2% of the community structure ( Figure 2). Ms. gryphiswaldense synthesizes nanosized magnetosomes (Fe3O4) by active uptake and reduction of Fe 3+ through ferric reductase [33]. Mn concentration in the wastewater may have been affected by this bacterium to some extent, since Mn can be incorporated into magnetite crystals [34].…”