Formation of the Fe-S cluster of ferredoxin in lysed spinach chloroplasts. [Spinacia oleracea]

Journal Article · Tue Jan 01 00:00:00 EST 1991 · Plant Physiology; (United States)

DOI:https://doi.org/10.1104/pp.95.1.97· OSTI ID:5680958

Takahashi, Yasuhiro; Mitsui, Akira; Matsubara, Hiroshi ^[1]

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1. Osaka Univ. (Japan)

In vitro formation of the {sup 35}S-labeled Fe-S cluster of ferredoxin (Fd) has been achieved by incubating apo-Fd and ({sup 35}S)cysteine with osmotically lysed chloroplasts of spinach (Spinacia oleracea). Correct integration of the {sup 35}S-labeled Fe-S cluster into Fd was verified on the basis of the following: (a) Under nondenaturing conditions, {sup 35}S-labeled holo-Fd showed the same electrophoretic mobility as authentic holo-Fd; (b) {sup 35}S-labeled holo-Fd showed an ability to bind Fd-NADP{sup +} reductase; (c) the {sup 35}S-labeled moiety was removed from the Fd polypeptide by TCA treatment but not by 2-mercaptoethanol treatment; (d) externally added pea II apo-Fd was converted to {sup 35}S-labeled holo-Fd. This reconstitution was dependent on both ATP and light, and formation of the {sup 35}S-labeled Fe-S cluster was observed upon addition of ATP or when an ATP generation-system was constructed in the light. In contrast, ATP-consuming systems abolished the Fe-S cluster formation. A non-hydrolyzable ATP analog was unable to serve as an ATP substitute, indicating the requirement of ATP hydrolysis for cluster formation. GTP was able to substitute for ATP, but CTP and UTP were less effective. Fe-S cluster formation in lysed chloroplasts was stimulated by light even in the presence of added ATP. Light stimulation was inhibited by DCMU or methyl viologen but not by NH{sub 4}{sup +}. NADPH was able to substitute for light, indicating that light energy is required for the production of reducing compounds such as NADPH in addition to the generation of ATP.

• OSTI ID: 5680958
• Journal Information: Plant Physiology; (United States), Vol. 95:1; ISSN 0032-0889
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
FERREDOXIN
BIOCHEMICAL REACTION KINETICS
SPINACH
PHYSIOLOGY
ATP
CHLOROPLASTS
CYSTEINE
IN VITRO
NADP
SULFUR 35
TRACER TECHNIQUES
VISIBLE RADIATION
AMINO ACIDS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
CARBOXYLIC ACIDS
CELL CONSTITUENTS
COENZYMES
DAYS LIVING RADIOISOTOPES
ELECTROMAGNETIC RADIATION
EVEN-ODD NUCLEI
FOOD
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MAGNOLIOPHYTA
MAGNOLIOPSIDA
METALLOPROTEINS
NUCLEI
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PLANTS
PROTEINS
RADIATIONS
RADIOISOTOPES
REACTION KINETICS
SULFUR ISOTOPES
THIOLS
VEGETABLES
551001* - Physiological Systems- Tracer Techniques