Plant Molecular Biology 12: 463-474, 1989 © 1989 Kluwer Academic Publishers. Printed in Belgium
463
Isolation and characterization of cDNA clones encoding the 17.9 and 8.1 kDa subunits of Photosystem I from Chlamydomonas reinhardtii Lars-Gunnar Franz6n, 1. Gerhard Frank, 2 Herbert Zuber 2 and Jean-David Rochaix 1 1Departments of Molecular Biology and Plant Biology, University of Geneva, Geneva, Switzerland; 2Institute of Molecular Biology and Biophysics, EidgenOssische Technische Hochschule, HiJnggerberg, Zfirich, Switzerland; * address for correspondence: L.-G. Franzdn, D~partement de Biologie Moldculaire, Universit~ de Gen~ve, 30 quai Ernest-Ansermet, CH-1211 Gen~ve 4, Switzerland Received 29 November 1988; accepted in revised form 2 February 1989
Key words: cDNA sequence, Chlamydomonas reinhardtii, chloroplast, nuclear-encoded subunits, Photosystem I, transit peptide
Abstract
cDNA clones encoding two Photosystem I subunits of Chlamydomonas reinhardtii with apparent molecular masses of 18 and 11 kDa (thylakoid polypeptides 21 and 30; P21 and P30 respectively) were isolated using oligonucleotides, the sequences of which were deduced from the N-terminal amino acid sequences of the proteins. The cDNAs were sequenced and used to probe Southern and Northern blots. The Southern blot analysis indicates that both proteins are encoded by single-copy genes. The mRNA sizes of the two components are 1400 and 740 nucleotides, respectively. Comparison between the open reading frames of the cDNAs and the N-terminal amino acid sequences of the proteins indicates that the molecular masses of the mature proteins are 17.9 (P21) and 8.1 kDa (P30). Analysis of the deduced protein sequences predicts that both subunits are extrinsic membrane proteins with net positive charges. The amino acid sequences of the transit peptides suggest that P21 and P30 are routed towards the lumenal and stromal sides of the thylakoid membranes, respectively.
Abbreviations: OEE1, 2 and 3, oxygen evolution enhancer proteins 1, 2 and 3; Rubisco, ribulose bisphosphate carboxylase/oxygenase; PS, photosystem; P21 and P30, C. reinhardtii thylakoid polypeptides 21 and 30
Introduction
In the photosynthetic electron transport chain, Photosystem I (PS I) catalyses the light-driven electron transfer from plastocyanin to ferredoxin.
The absorption of a light quantum results in an electron transfer from the reaction-centre chlorophyll P700 to a series of electron acceptors, namely Ao (probably a chlorophyll molecule), A 1 (probably a quinone) and the iron-sulphur centres
The nucleotide sequence data reported will appear in the EMBL, GenBank and DDBJ Nucleotide Sequence Databases under the accession numbers X13495 and X13496.
464 X, B and A. P700 is then re-reduced by plastocyanin and the electron on the P S I acceptor side is transferred to ferredoxin. P700 and the first acceptors Ao, A~ and FeS x appear to be located on two chloroplast-encoded large subunits (80-85kDa). The P S I core complex also contains several smaller subunits (
463
Isolation and characterization of cDNA clones encoding the 17.9 and 8.1 kDa subunits of Photosystem I from Chlamydomonas reinhardtii Lars-Gunnar Franz6n, 1. Gerhard Frank, 2 Herbert Zuber 2 and Jean-David Rochaix 1 1Departments of Molecular Biology and Plant Biology, University of Geneva, Geneva, Switzerland; 2Institute of Molecular Biology and Biophysics, EidgenOssische Technische Hochschule, HiJnggerberg, Zfirich, Switzerland; * address for correspondence: L.-G. Franzdn, D~partement de Biologie Moldculaire, Universit~ de Gen~ve, 30 quai Ernest-Ansermet, CH-1211 Gen~ve 4, Switzerland Received 29 November 1988; accepted in revised form 2 February 1989
Key words: cDNA sequence, Chlamydomonas reinhardtii, chloroplast, nuclear-encoded subunits, Photosystem I, transit peptide
Abstract
cDNA clones encoding two Photosystem I subunits of Chlamydomonas reinhardtii with apparent molecular masses of 18 and 11 kDa (thylakoid polypeptides 21 and 30; P21 and P30 respectively) were isolated using oligonucleotides, the sequences of which were deduced from the N-terminal amino acid sequences of the proteins. The cDNAs were sequenced and used to probe Southern and Northern blots. The Southern blot analysis indicates that both proteins are encoded by single-copy genes. The mRNA sizes of the two components are 1400 and 740 nucleotides, respectively. Comparison between the open reading frames of the cDNAs and the N-terminal amino acid sequences of the proteins indicates that the molecular masses of the mature proteins are 17.9 (P21) and 8.1 kDa (P30). Analysis of the deduced protein sequences predicts that both subunits are extrinsic membrane proteins with net positive charges. The amino acid sequences of the transit peptides suggest that P21 and P30 are routed towards the lumenal and stromal sides of the thylakoid membranes, respectively.
Abbreviations: OEE1, 2 and 3, oxygen evolution enhancer proteins 1, 2 and 3; Rubisco, ribulose bisphosphate carboxylase/oxygenase; PS, photosystem; P21 and P30, C. reinhardtii thylakoid polypeptides 21 and 30
Introduction
In the photosynthetic electron transport chain, Photosystem I (PS I) catalyses the light-driven electron transfer from plastocyanin to ferredoxin.
The absorption of a light quantum results in an electron transfer from the reaction-centre chlorophyll P700 to a series of electron acceptors, namely Ao (probably a chlorophyll molecule), A 1 (probably a quinone) and the iron-sulphur centres
The nucleotide sequence data reported will appear in the EMBL, GenBank and DDBJ Nucleotide Sequence Databases under the accession numbers X13495 and X13496.
464 X, B and A. P700 is then re-reduced by plastocyanin and the electron on the P S I acceptor side is transferred to ferredoxin. P700 and the first acceptors Ao, A~ and FeS x appear to be located on two chloroplast-encoded large subunits (80-85kDa). The P S I core complex also contains several smaller subunits (
