Journal
BIOPHYSICAL JOURNAL
Volume 120, Issue 23, Pages 5395-5407Publisher
CELL PRESS
DOI: 10.1016/j.bpj.2021.10.023
Keywords
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Categories
Funding
- Fundacao para a Ciencia e a Tecnologia (FCT) [SFRH/BD/145039/2019, SFRH/BPD/114848/2016, PTDC/BIA-BQM/31981/2017, PTDC/BIA-BQM/4967/2020]
- Applied Molecular Biosciences Unit -UCIBIO - FCT [UIDB/04378/2020, UIDP/04378/2020]
- Associate Laboratory Institute for Health and Bioeconomy i4HB [LA/P/0140/2020]
- Instituto de Tecnologia Quimica e Biologica Antonio Xavier -Molecular, Structural and Cellular Microbiology Research Unit (ITQB-MOSTMICRO) [UIDB/04612/2020, UIDP/04612/2020]
- FCT
- Fundo Europeu de Desenvolvimento Regional (FEDER) under the PT2020 Partnership Agreement
- European Union's Horizon 2020 research and innovation program [810856]
- FCT - FEDER through COMPETE 2020 [ROTEIRO/0031/2013, PINFRA/22161/2016]
- FCT - FEDER through Programa Operacional Ciencia e Inovacao (POCI) [ROTEIRO/0031/2013, PINFRA/22161/2016]
- FCT - FEDER through Programa Operacional Regional de Lisboa (PORL) [ROTEIRO/0031/2013, PINFRA/22161/2016]
- FCT through Programa de Investimentos e Despesas de Desenvolvimento da Administracao Central (PIDDAC)
- Fundação para a Ciência e a Tecnologia [PINFRA/22161/2016, SFRH/BPD/114848/2016, SFRH/BD/145039/2019] Funding Source: FCT
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Geobacter sulfurreducens has over 100 cytochromes, with the most abundant group being the PpcA family. A specific cytochrome, GSU0105, synthesized in Fe(III)-reducing conditions, has unique structural and heme coordination features compared to the PpcA family members. Through various biophysical techniques, GSU0105 was characterized to have distinct heme groups and a larger functional working redox potential range, indicating it as a member of a new group of triheme cytochromes.
Geobacter sulfurreducens possesses over 100 cytochromes that assure an effective electron transfer to the cell exterior. The most abundant group of cytochromes in this microorganism is the PpcA family, composed of five periplasmic triheme cytochromes with high structural homology and identical heme coordination (His-His). GSU0105 is a periplasmic triheme cytochrome synthetized by G. sulfurreducens in Fe(III)-reducing conditions but is not present in cultures grown on fumarate. This cytochrome has a low sequence identity with the PpcA family cytochromes and a different heme coordination, based on the analysis of its amino acid sequence. In this work, amino acid sequence analysis, site-directed mutagenesis, and complementary biophysical techniques, including ultraviolet-visible, circular dichroism, electron paramagnetic resonance, and nuclear magnetic resonance spectroscopies, were used to characterize GSU0105. The cytochrome has a low percentage of secondary structural elements, with features of alpha-helices and beta-sheets. Nuclear magnetic resonance shows that the protein contains three low-spin hemes (Fe(II), S = 0) in the reduced state. Electron paramagnetic resonance shows that, in the oxidized state, one of the hemes becomes high-spin (Fe(III), S = 5/2), whereas the two others remain low-spin (Fe(III), S = 1/2). The data obtained also indicate that the heme groups have distinct axial coordination. The apparent midpoint reduction potential of GSU0105 (-154 mV) is pH independent in the physiological range. However, the pH modulates the reduction potential of the heme that undergoes the low-to high-spin interconversion. The reduction potential values of cytochrome GSU0105 are more distinct compared to those of the PpcA family members, providing the protein with a larger functional working redox potential range. Overall, the results obtained, together with an amino acid sequence analysis of different multiheme cytochrome families, indicate that GSU0105 is a member of a new group of triheme cytochromes.
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