skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Measurement of heme efflux and heme content in isolated developing chloroplasts. [Cucumis sativus, cv. Sumter]

Abstract

Hemes destined for cytosolic hemoproteins must originate in one of the cellular compartments which have the capacity for heme synthesis, namely the chloroplast or the mitochondria. Since developing chloroplasts from greening cucumber (Cucumis sativus, cv. Sumter) cotyledons are known to contain complete heme and chlorophyll biosynthetic pathways, they were tested for their capacity export hemes. Picomole quantities of heme were measured by reconstitution of the heme with apo-peroxidase and subsequent determination of peroxidase activity. The assay method was sensitive (as little as 0.7 picomole of heme could be detected in a volume of 100 microliters) and was linear with heme concentration. When intact plastids were incubated with apo-peroxidase, a steady-state rate of efflux between 0.12 and 0.45 picomole heme/minute/milligram plastid protein was measured. The efflux rate was not due to plastid breakage and could be enhanced by incubating with the heme precursor, {delta}-aminolevulinic acid. Cold acetone extraction removed 47 {plus minus} 17 picomoles heme/milligram plastid protein from the total b-type heme pool in the chloroplasts (166 {plus minus} 9 picomoles heme/milligram protein, by acid-acetone extraction). The reconstitution technique provided a similar estimate of readily exchangeable heme in the plastid, 37 {plus minus} 8 picomoles heme/milligram protein (or 6 micromolar inmore » the plastids). These values may be indicative of a free heme pool which exists in the chloroplast.« less

Authors:
;  [1]
  1. (Clemson Univ., SC (USA))
Publication Date:
OSTI Identifier:
5626595
Resource Type:
Journal Article
Resource Relation:
Journal Name: Plant Physiology; (USA); Journal Volume: 94:3
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; CUCUMBERS; PHYSIOLOGY; HEME; MEASURING METHODS; BIOLOGICAL PATHWAYS; CHLOROPLASTS; ENZYME ACTIVITY; PEROXIDASES; CARBOXYLIC ACIDS; CELL CONSTITUENTS; ENZYMES; FOOD; HETEROCYCLIC ACIDS; HETEROCYCLIC COMPOUNDS; MAGNOLIOPHYTA; MAGNOLIOPSIDA; ORGANIC ACIDS; ORGANIC COMPOUNDS; ORGANIC NITROGEN COMPOUNDS; OXIDOREDUCTASES; PIGMENTS; PLANTS; PORPHYRINS; VEGETABLES; 551000* - Physiological Systems

Citation Formats

Thomas, J., and Weinstein, J.D. Measurement of heme efflux and heme content in isolated developing chloroplasts. [Cucumis sativus, cv. Sumter]. United States: N. p., 1990. Web. doi:10.1104/pp.94.3.1414.
Thomas, J., & Weinstein, J.D. Measurement of heme efflux and heme content in isolated developing chloroplasts. [Cucumis sativus, cv. Sumter]. United States. doi:10.1104/pp.94.3.1414.
Thomas, J., and Weinstein, J.D. Thu . "Measurement of heme efflux and heme content in isolated developing chloroplasts. [Cucumis sativus, cv. Sumter]". United States. doi:10.1104/pp.94.3.1414.
@article{osti_5626595,
title = {Measurement of heme efflux and heme content in isolated developing chloroplasts. [Cucumis sativus, cv. Sumter]},
author = {Thomas, J. and Weinstein, J.D.},
abstractNote = {Hemes destined for cytosolic hemoproteins must originate in one of the cellular compartments which have the capacity for heme synthesis, namely the chloroplast or the mitochondria. Since developing chloroplasts from greening cucumber (Cucumis sativus, cv. Sumter) cotyledons are known to contain complete heme and chlorophyll biosynthetic pathways, they were tested for their capacity export hemes. Picomole quantities of heme were measured by reconstitution of the heme with apo-peroxidase and subsequent determination of peroxidase activity. The assay method was sensitive (as little as 0.7 picomole of heme could be detected in a volume of 100 microliters) and was linear with heme concentration. When intact plastids were incubated with apo-peroxidase, a steady-state rate of efflux between 0.12 and 0.45 picomole heme/minute/milligram plastid protein was measured. The efflux rate was not due to plastid breakage and could be enhanced by incubating with the heme precursor, {delta}-aminolevulinic acid. Cold acetone extraction removed 47 {plus minus} 17 picomoles heme/milligram plastid protein from the total b-type heme pool in the chloroplasts (166 {plus minus} 9 picomoles heme/milligram protein, by acid-acetone extraction). The reconstitution technique provided a similar estimate of readily exchangeable heme in the plastid, 37 {plus minus} 8 picomoles heme/milligram protein (or 6 micromolar in the plastids). These values may be indicative of a free heme pool which exists in the chloroplast.},
doi = {10.1104/pp.94.3.1414},
journal = {Plant Physiology; (USA)},
number = ,
volume = 94:3,
place = {United States},
year = {Thu Nov 01 00:00:00 EST 1990},
month = {Thu Nov 01 00:00:00 EST 1990}
}
  • The gibberellin (GA) binding properties of a cytosol fraction from hypocotyls of cucumber (Cucumis sativus L. cv National Pickling) were examined using a DEAE filter paper assay, (/sup 3/H)GA/sub 4/, and over 20 GAs, GA derivatives and other growth regulators. The results demonstrate structural specificity of the binding protein for ..gamma..-lactonic C-19 GAs with a 3 ..beta..-hydroxyl and a C-6 carboxyl group. Additional hydroxylations of the A, C, or D ring of the ent-gibberellane skeleton and methylation of the C-6 carboxyl impede or abolish binding affinity. Bioassay data are generally supported by the in vitro results but significantly GA/sub 9/more » and GA/sub 36/, both considered to be precursors of GA/sub 4/ in cucumber, show no affinity for the binding protein. The results are discussed in relation to the active site of the putative GA/sub 4/ receptor in cucumber.« less
  • The effect of carbon monoxide and light on the respiration of a number of plant tissues were examined. The respiration of root or other tissue was measured at 25/sup 0/C by standard manometric techniques in a ratio of 95% CO and 5% O/sub 2/. The respiration of all eleven tissues studied was strongly inhibited by carbon monoxide. In ten of the eleven cases examined the inhibition was largely or completely eliminated by irradiation of the tissue with light. The evidence fairly well precludes the participation of a tyrosinase and definitely supports the participation of a cytochrome oxidase in respiration. 5more » references, 1 table.« less
  • Carbon exchange capacity of cucumber (Cucumis sativus L.) germinated and grown in controlled environment chambers at 1000 microliters per liter CO/sub 2/ decreased from the vegetative growth stage to the fruiting stage, during which time capacity of plants grown at 350 microliters per liter increased. Carbon exchange rates (CERs) measured under growth conditions during the fruiting period were, in fact, lower in plants grown at 1000 microliters per liter CO/sub 2/ than those grown at 350. Progressive decreases in CERs in 1000 microliters per liter plants were associated with decreasing stomatal conductances and activities of ribulose bisphosphate carboxylase and carbonicmore » anhydrase. Leaf starch concentrations were higher in 1000 microliters per liter CO/sub 2/ grown-plants than in 350 microliters per liter grown plants but calcium and nitrogen concentrations were lower, the greatest difference occurring at flowering. Sucrose synthase and sucrose-P-synthase activities were similar in 1000 microliters per liter compared to 350 microliters per liter plants during vegetative growth and flowering but higher in 350 microliters per liter plants at fruiting. The decreased carbon exchange rates observed in this cultivar at 1000 microliters per liter CO/sub 2/ could explain the lack of any yield increase when compared with plants grown at 350 microliters per liter.« less
  • Two cucumber and eight lettuce varieties were grown in two air-supported, closed-environment plastic greenhouses, one with approximately ambient CO2 levels, and the other enriched with 1400 ppm CO2. Diesel exhaust gas was the source of the carbon dioxide. Once the exhaust gases were scrubbed through seawater and put through an activated charcoal filter, essentially no other gases entered the greenhouse along with the CO2. Cucumbers grown in the enriched environment came into production one week earlier, and one variety produced significantly higher yields, than those grown at near ambient levels of CO2. Lettuce grown in the CO2 enriched greenhouse weighed,more » at market maturity, nearly twice as much as lettuce grown at ambient levels.« less