A Secondary Processing Site in the Precursor of the Small Subunit of Ribulose Bisphosphate Carboxylase of <i>Chlamydomonas reinhardtii</i> y-1

Marks, Dawn B.; Keller, Barbara; Hoober, J. Kenneth

doi:10.1104/pp.81.2.702

Cited by 10 publications

(6 citation statements)

References 8 publications

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“…Apparently CAB precursors fold in vitro into a set of conformations, among which only a fraction can be processed. The partial processing of CAB precursors was in marked contrast to the complete processing in vitro of the precursor to the small subunit of Rubisco, a soluble stromal protein (Marks et al 1986). The efficient processing of CAB precursors in vivo implies that additional factors such as molecular chaperones or membranes are involved inside the cell, which maintain the precursors in a state amenable to processing.…”

Section: B Kinetics Of Accumulation Of Cab Proteinsmentioning

confidence: 90%

“…Luminal proteins are synthesized as precursors with a bipartite N-terminal leader sequence. The proximal segment of the leader serves as the transit sequence that enables entry of the protein into the stroma, where this segment is removed by a specific peptidase (Robinson and Ellis 1984, Marks et al 1986). The distal, more hydrophobic segment--a typical'signal' sequence, is required for transfer of the protein across thylakoid membranes.…”

Section: B Proteinsmentioning

confidence: 99%

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Biogenesis of thylakoid membranes with emphasis on the process in Chlamydomonas

et al. 1994

Photosynth Res

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Recent results obtained by electron microscopic and biochemical analyses of greening Chlamydomonas reinhardtii y1 suggest that localized expansion of the plastid envelope is involved in thylakoid biogenesis. Kinetic analyses of the assembly of light-harvesting complexes and development of photosynthetic function when degreened cells of the alga are exposed to light suggest that proteins integrate into membrane at the level of the envelope. Current information, therefore, supports the earlier conclussion that the chloroplast envelope is a major biogenic structure, from which thylakoid membranes emerge. Chloroplast development in Chlamydomonas provides unique opportunities to examine in detail the biogenesis of thylakoids.

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Section: B Kinetics Of Accumulation Of Cab Proteinsmentioning

confidence: 90%

Section: B Proteinsmentioning

confidence: 99%

Biogenesis of thylakoid membranes with emphasis on the process in Chlamydomonas

et al. 1994

Photosynth Res

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“…If pSS from pea was previously treated with iodoacetate, the intermediate was the only proteolytic product after incubation with pea peptidase, which in vitro nonnally processed pSS of pea to the mature form. When pSS from C. reinhardii was bound to antibodies and treated with a soluble cell extract, it was cleaved to the mature form via an intermediate protein (Marks et al, 1986). Therefore, pSS from C. reinhardii was also claimed to be processed in two steps.…”

Section: Discussionmentioning

confidence: 99%

“…Not only proteins of the thylakoid membranes, e.g. apoproteins of the light-harvesting complex I1 (LHCII) (Clark et al, 1989;Clark and Lamppa, 1992), but also proteins located in the stroma, such as the small subunit of ribulose-1,5-bisphosphate carboxylase (SS) or ribosomal proteins (Robinson and Ellis, 1984b;Marks et al, 1986, Mishkind et al, 1985Schmidt et al, 1985) were claimed to be processed via an intermediate-sized form. On the physiological significance of such a two-step mechanism and of the intermediate-sized form of the import protein only speculations are possible.…”

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Partial purification and properties of enzymes involved in the processing of a chloroplast import protein from Chlamydomonas reinhardii

Su¹,

Boschetti²

1993

European Journal of Biochemistry

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Two stromal peptidases (SPP‐1 and SPP‐2) were partially purified from chloroplasts of Chlamydomonas reinhardii. They specifically processed in vitro the precursor of the small subunit of ribulose‐1,5‐bisphosphate carboxylase (pSS), which had been synthesized by using the cloned rbcS‐2 gene of Chlamydomonas. SPP‐1 shortened pSS to an intermediate‐sized form (iSS), while SPP‐2 cut pSS and iSS to the mature small subunit SS. N‐terminal amino acid sequencing demonstrated that the reaction product obtained with SPP‐2 had an N‐terminus identical to natural SS, and that iSS derived from pSS by hydrolysis at the amino side of the methionine located within the transit sequence. By gel filtration, apparent molecular masses of 340 kDa and 90 kDa were determined for SPP‐1 and SPP‐2, respectively. The comparison of these molecular masses with the protein patterns obtained by SDS/PAGE of the partially purified enzymes suggested that at least SPP‐1 was a multimeric protein. The enzymes differed also in their pH optima of about 8 (SPP‐1) and 9 (SPP‐2) and in their sensitivity to different inhibitors. However, both enzymes seem to be serine proteases as they were completely blocked by N‐α‐tosyl‐l‐lysinechloromethane or tosylphenylalaninechloromethane, respectively. Competition experiments, using either mature SS or a synthetic hexadecapeptide with 15 amino acids similar to the C‐terminal end of the transit sequence of pSS, indicated that SPP‐2 had some affinities not only to the transit sequence of pSS, but especially to sequences in the mature protein part. We conclude that SPP‐2 in Chlamydomonas is the enzyme involved in import of pSS into chloroplasts and responsible for its processing by a one‐step mechanism.

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“…Two reports [2,11 ] have demonstrated SPP activity in Chlamydomonas reinhardtii, but this activity was not characterised in detail. The little information available has suggested that the processing mechanisms in this alga may be significantly different from those in higher plants.…”

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confidence: 99%

The stromal processing peptidase activities from Chlamydomonas reinhardtii and Pisum sativum: unexpected similarities in reaction specificity

1993

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We have partially purified the stromal processing peptidase from Chlamydomonas reinhardtii and compared the properties of this activity with those of the pea counterpart. Whereas previous studies have suggested that the two enzymes may have significantly different reaction specificities, we find that they are in fact very similar. Both enzymes process precursors of two higher-plant thylakoid lumen proteins, and one C. reinhardtii lumenal protein, to similar intermediate-size forms. However, whereas the algal enzyme processes the precursor of C. reinhardtii Rubisco small subunit to the correct mature size, this precursor is cleaved only to an intermediate size by the pea enzyme. The small subunit precursor from pea appears to be cleaved by both enzymes in a similar manner. In terms of sensitivity to inhibitors, the two activities are notably different; the pea enzyme has previously been shown to be inhibited by several types of heavy-metal chelator, but we have found that none of these compounds affect the algal activity.

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A Secondary Processing Site in the Precursor of the Small Subunit of Ribulose Bisphosphate Carboxylase of Chlamydomonas reinhardtii y-1

Cited by 10 publications

References 8 publications

Biogenesis of thylakoid membranes with emphasis on the process in Chlamydomonas

Biogenesis of thylakoid membranes with emphasis on the process in Chlamydomonas

Partial purification and properties of enzymes involved in the processing of a chloroplast import protein from Chlamydomonas reinhardii

The stromal processing peptidase activities from Chlamydomonas reinhardtii and Pisum sativum: unexpected similarities in reaction specificity

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