{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,13]],"date-time":"2026-10-13T19:38:13Z","timestamp":1791920293774,"version":"4.3.4"},"reference-count":34,"publisher":"Springer Science and Business Media LLC","issue":"1-2","license":[{"start":{"date-parts":[[2021,5,1]],"date-time":"2021-05-01T00:00:00Z","timestamp":1619827200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2021,5,2]],"date-time":"2021-05-02T00:00:00Z","timestamp":1619913600000},"content-version":"vor","delay-in-days":1,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/501100009027","name":"Advanced Low Carbon Technology Research and Development Program","doi-asserted-by":"publisher","award":["JPMJAL1402"],"award-info":[{"award-number":["JPMJAL1402"]}],"id":[{"id":"10.13039\/501100009027","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["18J20176"],"award-info":[{"award-number":["18J20176"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["20J00105"],"award-info":[{"award-number":["20J00105"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Photosynth Res"],"published-print":{"date-parts":[[2021,5]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>In photosynthetic organisms, it is recognized that the intracellular redox ratio of NADPH is regulated within an appropriate range for the cooperative function of a wide variety of physiological processes. However, despite its importance, there is large variability in the values of the NADPH fraction [NADPH\/(NADPH\u2009+\u2009NADP<jats:sup>+<\/jats:sup>)] quantitatively estimated to date. In the present study, the light response of the NADPH fraction was investigated by applying a novel NADP(H) extraction method using phenol \/ chloroform \/ isoamyl alcohol (PCI) in the cyanobacterium <jats:italic>Synechocystis<\/jats:italic> sp. PCC 6803. The light response of NADP(H) observed using PCI extraction was qualitatively consistent with the NAD(P)H fluorescence time course measured in vivo. Moreover, the results obtained by PCI extraction and the fluorescence-based methods were also consistent in a mutant lacking the ability to oxidize NAD(P)H in the respiratory chain, and exhibiting a unique NADPH light response. These observations indicate that the PCI extraction method allowed quantitative determination of NADP(H) redox. Notably, the PCI extraction method showed that not all NADP(H) was oxidized or reduced by light\u2013dark transition. Specifically, the fraction of NADPH was 42% in the dark-adapted cell, and saturated at 68% in light conditions.<\/jats:p>","DOI":"10.1007\/s11120-021-00835-1","type":"journal-article","created":{"date-parts":[[2021,5,3]],"date-time":"2021-05-03T15:29:46Z","timestamp":1620055786000},"page":"57-66","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":24,"title":["Quantification of NAD(P)H in cyanobacterial cells by a phenol extraction method"],"prefix":"10.1007","volume":"148","author":[{"given":"Kenya","family":"Tanaka","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ginga","family":"Shimakawa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hiro","family":"Tabata","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shoko","family":"Kusama","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chikahiro","family":"Miyake","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shuji","family":"Nakanishi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2021,5,2]]},"reference":[{"key":"835_CR1","doi-asserted-by":"publisher","first-page":"2219","DOI":"10.1021\/bi00511a023","volume":"20","author":"MR Badger","year":"1981","unstructured":"Badger MR, Lorimer GH (1981) Interaction of sugar phosphates with the catalytic site of ribulose-1,5-bisphosphate carboxylase. Biochemistry 20:2219\u20132225","journal-title":"Biochemistry"},{"key":"835_CR2","doi-asserted-by":"publisher","first-page":"4251","DOI":"10.1128\/JB.183.14.4251-4258.2001","volume":"183","author":"JW Cooley","year":"2001","unstructured":"Cooley JW, Vermaas WF (2001) Succinate dehydrogenase and other respiratory pathways in thylakoid membranes of Synechocystis sp. strain PCC 6803: capacity comparisons and physiological function. J Bacteriol 183:4251\u20134258","journal-title":"J Bacteriol"},{"key":"835_CR3","doi-asserted-by":"publisher","first-page":"1617","DOI":"10.1016\/0006-291X(72)90527-X","volume":"49","author":"LH Grimme","year":"1972","unstructured":"Grimme LH, Boardman NK (1972) Photochemical activities of a particle fraction P 1 obtained rom the green alga Chlorella fusca. Biochem Biophys Res Commun 49:1617\u20131623","journal-title":"Biochem Biophys Res Commun"},{"key":"835_CR4","doi-asserted-by":"publisher","first-page":"11","DOI":"10.1093\/pcp\/pcm158","volume":"49","author":"S Hishiya","year":"2008","unstructured":"Hishiya S, Hatakeyama W, Mizota Y, Hosoya-Matsuda N, Motohashi K, Ikeuchi M, Hisabori T (2008) Binary reducing equivalent pathways using NADPH-thioredoxin reductase and ferredoxin-thioredoxin reductase in the cyanobacterium Synechocystis sp. strain PCC 6803. Plant Cell Physiol 49:11\u201318","journal-title":"Plant Cell Physiol"},{"key":"835_CR5","doi-asserted-by":"publisher","first-page":"355","DOI":"10.1016\/j.bbabio.2014.12.001","volume":"1847","author":"SC Holland","year":"2015","unstructured":"Holland SC, Kappell AD, Burnap RL (2015) Redox changes accompanying inorganic carbon limitation in Synechocystis sp. PCC 6803. Biochim Biophys Acta 1847:355\u2013363","journal-title":"Biochim Biophys Acta"},{"key":"835_CR6","doi-asserted-by":"publisher","first-page":"105","DOI":"10.1016\/j.jplph.2016.09.002","volume":"205","author":"Y Ishikawa","year":"2016","unstructured":"Ishikawa Y, Miyagi A, Haishima Y, Ishikawa T, Nagano M, Yamaguchi M, Hihara Y, Kawai-Yamada M (2016) Metabolomic analysis of NAD kinase-deficient mutants of the cyanobacterium Synechocystis sp. PCC 6803. J Plant Physiol 205:105\u2013112","journal-title":"J Plant Physiol"},{"key":"835_CR7","doi-asserted-by":"publisher","first-page":"847","DOI":"10.3389\/fpls.2019.00847","volume":"10","author":"Y Ishikawa","year":"2019","unstructured":"Ishikawa Y, Kawai-Yamada M (2019) Physiological significance of NAD kinases in cyanobacteria. Front Plant Sci 10:847","journal-title":"Front Plant Sci"},{"key":"835_CR8","doi-asserted-by":"publisher","first-page":"654","DOI":"10.1111\/tpj.14262","volume":"98","author":"Y Ishikawa","year":"2019","unstructured":"Ishikawa Y, Miyagi A, Ishikawa T, Nagano M, Yamaguchi M, Hihara Y, Kaneko Y, Kawai-Yamada M (2019) One of the NAD kinases, sll1415, is required for the glucose metabolism of Synechocystis sp. PCC 6803. Plant J 98:654\u2013666","journal-title":"Plant J"},{"key":"835_CR9","doi-asserted-by":"publisher","first-page":"1309","DOI":"10.1042\/BCJ20200038","volume":"477","author":"S Ito","year":"2020","unstructured":"Ito S, Osanai T (2020) Unconventional biochemical regulation of the oxidative pentose phosphate pathway in the model cyanobacterium Synechocystis sp. PCC 6803. Biochem J 477:1309\u20131321","journal-title":"Biochem J"},{"key":"835_CR10","doi-asserted-by":"publisher","first-page":"792","DOI":"10.1016\/j.bbabio.2014.01.009","volume":"1837","author":"J Kauny","year":"2014","unstructured":"Kauny J, S\u00e9tif P (2014) NADPH fluorescence in the cyanobacterium Synechocystis sp. PCC 6803: a versatile probe for in vivo measurements of rates, yields and pools. Biochim Biophys Acta 1837:792\u2013801","journal-title":"Biochim Biophys Acta"},{"key":"835_CR11","doi-asserted-by":"publisher","first-page":"311","DOI":"10.1007\/978-1-4939-0473-0_26","volume":"1149","author":"SE Kern","year":"2014","unstructured":"Kern SE, Price-Whelan A, Newman DK (2014) Extraction and measurement of NAD(P)(+) and NAD(P)H. Methods Mol Biol 1149:311\u2013323","journal-title":"Methods Mol Biol"},{"key":"835_CR12","doi-asserted-by":"publisher","first-page":"367","DOI":"10.1002\/1873-3468.13605","volume":"594","author":"S Khorobrykh","year":"2020","unstructured":"Khorobrykh S, Tsurumaki T, Tanaka K, Tyystj\u00e4rvi T, Tyystj\u00e4rvi E (2020) Measurement of the redox state of the plastoquinone pool in cyanobacteria. FEBS Lett 594:367\u2013375","journal-title":"FEBS Lett"},{"key":"835_CR13","doi-asserted-by":"publisher","first-page":"2126","DOI":"10.1002\/2211-5463.12752","volume":"9","author":"Kimata-Ariga","year":"2019","unstructured":"Kimata-Ariga, Chikuma Y, Saitoh T, Miyata M, Yanagihara Y, Yamane K, Hase T (2019) NADP(H) allosterically regulates the interaction between ferredoxin and ferredoxin-NADP+ reductase. FEBS Open Bio 9:2126\u20132136","journal-title":"FEBS Open Bio"},{"key":"835_CR14","doi-asserted-by":"publisher","first-page":"1669","DOI":"10.1016\/j.bbabio.2006.08.004","volume":"1757","author":"J Kruk","year":"2006","unstructured":"Kruk J, Karpinski S (2006) An HPLC-based method of estimation of the total redox state of plastoquinone in chloroplasts, the size of the photochemically active plastoquinone-pool and its redox state in thylakoids of Arabidopsis. Biochim Biophys Acta 1757:1669\u20131675","journal-title":"Biochim Biophys Acta"},{"key":"835_CR15","doi-asserted-by":"publisher","first-page":"311","DOI":"10.1016\/S0005-2728(00)00198-5","volume":"1460","author":"G Latouche","year":"2000","unstructured":"Latouche G, Cerovic ZG, Montagnini F, Moya I (2000) Light-induced changes of NADPH fluorescence in isolated chloroplasts: a spectral and fluorescence lifetime study. Biochim Biophys Acta 1460:311\u2013329","journal-title":"Biochim Biophys Acta"},{"key":"835_CR16","doi-asserted-by":"publisher","first-page":"260","DOI":"10.1016\/0005-2728(75)90040-7","volume":"396","author":"K Lendzian","year":"1975","unstructured":"Lendzian K, Bassham JA (1975) Regulation of glucose-6-phosphate dehydrogenase in spinach chloroplasts by ribulose 1,5-diphosphate and NADPH\/NADP+ ratios. Biochim Biophys Acta 396:260\u2013275","journal-title":"Biochim Biophys Acta"},{"key":"835_CR17","doi-asserted-by":"publisher","first-page":"3238","DOI":"10.1038\/s41467-020-17056-0","volume":"11","author":"SL Lim","year":"2020","unstructured":"Lim SL, Voon CP, Guan X, Yang Y, Gardestr\u00f6m P, Lim BL (2020) In planta study of photosynthesis and photorespiration using NADPH and NADH\/NAD+ fluorescent protein sensors. Nat Commun 11:3238","journal-title":"Nat Commun"},{"key":"835_CR18","doi-asserted-by":"publisher","first-page":"1129","DOI":"10.1093\/pcp\/pcd038","volume":"41","author":"H Mi","year":"2000","unstructured":"Mi H, Klughammer C, Schreiber U (2000) Light-induced dynamic changes of NADPH fluorescence in Synechocystis PCC 6803 and its ndhB-defective mutant M55. Plant Cell Physiol 41:1129\u20131135","journal-title":"Plant Cell Physiol"},{"key":"835_CR19","doi-asserted-by":"publisher","first-page":"99","DOI":"10.1111\/j.1432-1033.1992.tb19833.x","volume":"203","author":"MI Muro-Pastor","year":"1992","unstructured":"Muro-Pastor MI, Florencio FJ (1992) Purification and properties of NADP-isocitrate dehydrogenase from the unicellular cyanobacterium Synechocystis sp. PCC 6803. Eur J Biochem 203:99\u2013105","journal-title":"Eur J Biochem"},{"key":"835_CR20","doi-asserted-by":"publisher","first-page":"31630","DOI":"10.1074\/jbc.M003706200","volume":"275","author":"H Ohkawa","year":"2000","unstructured":"Ohkawa H, Pakrasi HB, Ogawa T (2000) Two types of functionally distinct NAD(P)H dehydrogenases in Synechocystis sp. strain PCC6803. J Biol Chem 275:31630\u201331634","journal-title":"J Biol Chem"},{"key":"835_CR21","doi-asserted-by":"publisher","first-page":"512","DOI":"10.1111\/1462-2920.12170","volume":"16","author":"Osanai","year":"2014","unstructured":"Osanai, Oikawa A, Shirai T, Kuwahara A, Iijima H, Tanaka K, Ikeuchi M, Kondo A, Saito K, Hirai MY (2014) Capillary electrophoresis-mass spectrometry reveals the distribution of carbon metabolites during nitrogen starvation in Synechocystis sp. PCC 6803. Environ Microbiol 16:512\u2013524","journal-title":"Environ Microbiol"},{"key":"835_CR22","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1023\/A:1022421515027","volume":"75","author":"CA Raines","year":"2003","unstructured":"Raines CA (2003) The Calvin cycle revisited. Photosynth Res 75:1\u201310","journal-title":"Photosynth Res"},{"key":"835_CR23","doi-asserted-by":"publisher","first-page":"148256","DOI":"10.1016\/j.bbabio.2020.148256","volume":"1861","author":"P S\u00e9tif","year":"2020","unstructured":"S\u00e9tif P, Shimakawa G, Krieger-Liszkay A, Miyake C (2020) Identification of the electron donor to flavodiiron proteins in Synechocystis sp. PCC 6803 by in vivo spectroscopy. Biochim Biophys Acta Bioenerg 1861:148256","journal-title":"Biochim Biophys Acta Bioenerg"},{"key":"835_CR24","first-page":"1443","volume":"57","author":"K Shaku","year":"2016","unstructured":"Shaku K, Shimakawa G, Hashiguchi M, Miyake C (2016) Reduction-induced suppression of electron flow (RISE) in the photosynthetic electron transport system of Synechococcus elongatus PCC 7942. Plant Cell Physiol 57:1443\u20131453","journal-title":"Plant Cell Physiol"},{"key":"835_CR25","doi-asserted-by":"publisher","first-page":"886","DOI":"10.3389\/fmicb.2018.00886","volume":"9","author":"G Shimakawa","year":"2018","unstructured":"Shimakawa G, Shaku K, Miyake C (2018) Reduction-induced suppression of electron flow (RISE) is relieved by non-ATP-consuming electron flow in Synechococcus elongatus PCC 7942. Front Microbiol 9:886","journal-title":"Front Microbiol"},{"key":"835_CR26","doi-asserted-by":"publisher","first-page":"16019","DOI":"10.1073\/pnas.1918919117","volume":"117","author":"K Sugiura","year":"2020","unstructured":"Sugiura K, Mihara S, Fu N, Hisabori T (2020) Real-time monitoring of the in vivo redox state transition using the ratiometric redox state sensor protein FROG\/B. Proc Natl Acad Sci USA 117:16019\u201316026","journal-title":"Proc Natl Acad Sci USA"},{"key":"835_CR27","doi-asserted-by":"publisher","first-page":"5514","DOI":"10.1073\/pnas.90.12.5514","volume":"90","author":"KH Suss","year":"1993","unstructured":"Suss KH, Arkona C, Manteuffel R, Adler K (1993) Calvin cycle multienzyme complexes are bound to chloroplast thylakoid membranes of higher plants in situ. Proc Natl Acad Sci USA 90:5514\u20135518","journal-title":"Proc Natl Acad Sci USA"},{"key":"835_CR28","doi-asserted-by":"publisher","first-page":"1387","DOI":"10.1104\/pp.107.4.1387","volume":"107","author":"KH Suss","year":"1995","unstructured":"Suss KH, Prokhorenko I, Adler K (1995) In situ association of Calvin cycle enzymes, ribulose-1,5-bisphosphate carboxylase\/oxygenase activase, ferredoxin-NADP+ reductase, and nitrite reductase with thylakoid and pyrenoid membranes of Chlamydomonas reinhardtii chloroplasts as revealed by immunoelectron microscopy. Plant Physiol 107:1387\u20131397","journal-title":"Plant Physiol"},{"key":"835_CR29","doi-asserted-by":"publisher","first-page":"3009","DOI":"10.1093\/jxb\/ern157","volume":"59","author":"H Takahashi","year":"2008","unstructured":"Takahashi H, Uchimiya H, Hihara Y (2008) Difference in metabolite levels between photoautotrophic and photomixotrophic cultures of Synechocystis sp. PCC 6803 examined by capillary electrophoresis electrospray ionization mass spectrometry. J Exp Bot 59:3009\u20133018","journal-title":"J Exp Bot"},{"key":"835_CR30","doi-asserted-by":"publisher","first-page":"504","DOI":"10.1111\/j.1365-313X.2005.02391.x","volume":"42","author":"M Tamoi","year":"2005","unstructured":"Tamoi M, Miyazaki T, Fukamizo T, Shigeoka S (2005) The Calvin cycle in cyanobacteria is regulated by CP12 via the NAD(H)\/NADP(H) ratio under light\/dark conditions. Plant J 42:504\u2013513","journal-title":"Plant J"},{"key":"835_CR31","doi-asserted-by":"publisher","first-page":"1865","DOI":"10.1042\/BCJ20190124","volume":"477","author":"L Vogelsang","year":"2020","unstructured":"Vogelsang L, Dietz KJ (2020) Regulatory thiol oxidation in chloroplast metabolism, oxidative stress response and environmental signaling in plants. Biochem J 477:1865\u20131878","journal-title":"Biochem J"},{"key":"835_CR32","doi-asserted-by":"publisher","first-page":"10","DOI":"10.1111\/j.1399-3054.1993.tb01754.x","volume":"88","author":"B Wigge","year":"1993","unstructured":"Wigge B, Kr\u00f6mer S, Gardestr\u00f6m P (1993) The redox levels and subcellular distribution of pyridine nucleotides in illuminated barley leaf protoplasts studied by rapid fractionation. Physiol Plant 88:10\u201318","journal-title":"Physiol Plant"},{"key":"835_CR33","doi-asserted-by":"publisher","first-page":"E3967","DOI":"10.1073\/pnas.1604101113","volume":"113","author":"K Yoshida","year":"2016","unstructured":"Yoshida K, Hisabori T (2016) Two distinct redox cascades cooperatively regulate chloroplast functions and sustain plant viability. Proc Natl Acad Sci USA 113:E3967-3976","journal-title":"Proc Natl Acad Sci USA"},{"key":"835_CR34","doi-asserted-by":"publisher","first-page":"1551","DOI":"10.1126\/science.1210858","volume":"334","author":"S Zhang","year":"2011","unstructured":"Zhang S, Bryant DA (2011) The tricarboxylic acid cycle in cyanobacteria. Science 334:1551\u20131553","journal-title":"Science"}],"container-title":["Photosynthesis Research"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1007\/s11120-021-00835-1.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1007\/s11120-021-00835-1\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1007\/s11120-021-00835-1.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,5,26]],"date-time":"2021-05-26T12:14:58Z","timestamp":1622031298000},"score":1,"resource":{"primary":{"URL":"https:\/\/link.springer.com\/10.1007\/s11120-021-00835-1"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,5]]},"references-count":34,"journal-issue":{"issue":"1-2","published-print":{"date-parts":[[2021,5]]}},"alternative-id":["835"],"URL":"https:\/\/doi.org\/10.1007\/s11120-021-00835-1","relation":{},"ISSN":["0166-8595","1573-5079"],"issn-type":[{"value":"0166-8595","type":"print"},{"value":"1573-5079","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,5]]},"assertion":[{"value":"25 December 2020","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"12 April 2021","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"2 May 2021","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Declarations"}},{"value":"The authors declare that they have no conflict of interest.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Conflicts of interest"}}]}}