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A mutant of Hordeum vulgare L. (LaPr 85/84) deficient in serine: glyoxylate aminotransferase (EC 2.6.1.45) activity has been isolated. The plant also lacks serine: pyruvate aminotransferase and asparagine: glyoxylate aminotransferase activities. Genetic analysis of the mutation strongly indicates that these three activities are all carried on the same enzyme protein. The mutant is incapable of normal rates of photosynthesis in air but can be maintained at 0.7% CO2. The rate of photosynthesis cannot be restored by supplying hydroxypyruvate, glycerate, glutamate or ammonium sulphate through the xylem stream. This photorespiratory mutant demonstrates convincingly that photorespiration still occurs under conditions in which photosynthesis becomes insensitive to oxygen levels. Two major peaks and one minor peak of serine: glyoxylate aminotransferase activity can be separated in extracts of leaves of wild-type barley by diethylaminoethyl-sephacel chromatography. All three peaks are missing from the mutant, LaPr 85/84. The mutant showed the expected rate (50%) of ammonia release during photorespiration but produced CO2 at twice the wild-type rate when it was fed [(14)C]glyoxylate. The large accumulation of serine detected in the mutant under photorespiratory conditions shows the importance of the enzyme activity in vivo. The effect of the mutation on transient changes in chlorophyll a fluorescence initiated by changing the atmospheric CO2 concentration are presented and the role of the enzyme activity under nonphotorespiratory conditions is discussed.

Manipulation of the CO2 concentration of the atmosphere allows the selection of photorespiratory mutants from populations of seeds treated with powerful mutagens such as sodium azide. So far, barley lines deficient in activity of phosphoglycolate phosphatase, catalase, the glycine to serine conversion, glutamine synthetase, glutamate synthase, 2-oxoglutarate uptake and serine: glyoxylate aminotransferase have been isolated. In addition one line of pea lacking glutamate synthase activity and one barley line containing reduced levels of Rubisco are available. The characteristics of these mutations are described and compared with similar mutants isolated from populations of Arabidopsis. As yet, no mutant lacking glutamine synthetase activity has been isolated from Arabidopsis and possible reasons for this difference between barley and Arabidopsis are discussed. The value of these mutant plants in the elucidation of the mechanism of photorespiration and its relationships with CO2 fixation and amino acid metabolism are highlighted.

A mutant of barley (Hordeum vulgare L.), LaPr 88/29, deficient in NADH-dependent hydroxypyruvate reductase (HPR) activity has been isolated. The activities of both NADH (5%) and NADPHdependent (19%) HPR were severely reduced in this mutant compared to the wild type. Although lacking an enzyme in the main carbon pathway of photorespiration, this mutant was capable of CO2 fixation rates equivalent to 75% of that of the wild type, in normal atmospheres and 50% 02. There and subsequently reduced to glycerate under the auspices of HPR.At atmospheric concentrations of CO2 greater than 0.2%, carbon is directed mainly through the carboxylase activity of ribulose bisphosphate carboxylase/oxygenase. This property was utilized by Somerville and Ogren (22) to identify and maintain conditional lethal mutants, with deficiencies in the photorespiratory pathway and consequently incapable of growth at ambient levels of CO2 (0.034%).Definitive studies of HPR activity in plants are hampered by the apparent multiplicity of enzymes showing this activity (12, 13). The central importance in photorespiration of the NADH-dependent peroxisomal enzyme HPR has been accepted for almost 20 years (10,11,29) and is an enzyme thought to be especially active in green leaves (25,29). Recently, a novel HPR activity has been isolated and purified from spinach that can utilize NADPH in preference to NADH as a cofactor (12). This form of HPR has been shown to be localized in the cytosol (14). The existence of glyoxylate reductase activity is also partially confusing.Although mutants deficient in some of the many steps of photorespiration have been isolated from populations of mutated seed of Arabidopsis thaliana (22) MATERIALS AND METHODS Plant MaterialThe M2 seeds of Hordeum vulgare L. (cv Klaxon) used for mutant screening were produced as described previously (20) in a glasshouse supplemented to a CO2 concentration of 0.7% (v/v). Selection of air-sensitive plants was carried out in a glasshouse with atmospheric CO2 concentrations, maintained at 200C. Preparation of Crude Enzyme ExtractThe preparation of crude enzyme extracts for use in measurements of enzyme activity and with PAGE was Plant Physiol. Vol. 91,1989 Assay of Hydroxypyruvate and Glyoxylate Reductase ActivityThe assays for NADH and NADPH-HPR activities were based on those described by Stabenau (24) and Kleczkowski and Randall (12). The final concentrations in 1 mL of assay mixture were 5 mm hydroxypyruvate, 25 mm phosphate buffer (pH 6.3), 0.2 mm NADH (NADPH) and 50 ,L crude enzyme extract. Reactions were initiated by the addition of hydroxypyruvate and the oxidation of NADH (NADPH) was monitored at 340 nm.GR activity was determined using the same assay system by the substitution of 5 mm glyoxylate for hydroxypyruvate in the reaction mixture. PAGENative PAGE and staining for NADH-HPR activity were carried out using 4.5% acrylamide slab gels (15, 27), other related reductases not being reversible are not detected by this method (12). Samples were loaded onto gels after mix...

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