Diss.ETHNr.7610 Der Einfluss des Aktivierungszustandes und der katalytischen Eigenschaften der Ribulose-1.5-bisphosphatCarboxylase/Oxygenase auf die Photosynthese intakter Blätter von Weissklee (Trifolium repens L.) ABHANDLUNG zur Erlangung des Titels eines Doktors der technischen Wissenschaften der EIDGENÖSSISCHEN TECHNISCHEN HOCHSCHULE ZÜRICH vorgelegt von JOHANNES SCHNYDER dipl. ing. agr. ETH geboren am 25. April 1954 von Diessbach bei Büren (BE) Angenommen auf Antrag von Prof. Dr. J. Nösberger, Referent Prof. Dr. Ph. Matile, Korreferent
- PR'/RPS') 59 V. ZUSAMMENFASSUNG Die Beziehung zwischen der Nettophotosynthese und den katalytischen Eigenschaften sowie der Aktivität der RuBPCO wurde bei intakten Blattern von Weissklee untersucht Die Nettophotosynthese und die Photorespiration wurden mit Infra rot-gaswechsel-messungen in einem offenen System und der **C/ 12C-Technik (20 Sekunden Markierdauer) gemessen Der Aktivie rungszustand der RuBPCO im Blatt wurde als substratgesattigte An fangsgeschwindigkeit der Carboxylaseaktivitat C02-frei bereiteter Blattextrakte bestimmt RuBP und ATP wurden enzymatisch gemessen Die Nettophotosynthese (NPS) kann mit der Beziehung NPS = RPS (1 beschrieben werden (RPS' = reelle Photo synthese, PR' = Photorespiration) Die katalytischen Eigenschaften der RuBPCO beeinflussen die Nettophotosynthese über das PR'/RPS' - Verhältnis Dieses nahm mit steigendem 02/C02-Verhaltnis in der Um gebungsluft der Blatter linear zu Steigende Messtemperatur forderte PR'/RPS' in intakten Blattern von Weissklee ebenfalls Ein geringer Teil dieser Zunahme war auf die, im Vergleich zu 02, stärkere Ab nahme der Loslichkeit von C02 mit steigender Temperatur zurückzufüh ren Der übrige Teil dieser Zunahme beruht auf einem unbekannten Effekt Die Zunahme des PR'/RPS'-Verhaltnisses mit steigender Mosstemperatur ist eine der Ursachen für die relativ schwache Temperaturabhangigkeit der Nettophotosynthese bei atmosphanschen 02- und C02-Konzentrationen Bei Blattern aus Kaltanzucht (10/7 C Tag-/ Nachttemperatur) wurde das PR'/RPS'-Verhaltnis, im Vergleich zu Blattern aus Warmanzucht (20/16 "C Tag-/Nachttemperatur) mit stei gender Messtemperatur starker gefordert Diese stärkere Zunahme erklart, mindestens zum Teil, die schwächere Temperaturabhangigkeit der Nettophotosynthese dieser Blatter Bei Blattern aus Warmanzucht zeigte die 02-Hemmung der Photosyn these eine stärkere Temperaturabhangigkeit als das PR'/RPS'-Verhaltnis, und die 02- und C02-Abhangigkeit der reellen Photosynthese
61 Summary The influence of the activation State of Ribulose-1.5-bisphosphate carboxylase/oxygenase (RuBPCO) and the catalytic properties of this enzyme on photosynthesis of intact leaves of white clover (Trifolium repens L.) was examined. The activation state in vivo of RuBPCO was estimated by measuring the initial carboxylase activity of rapidly prepared C02 free extracts of leaves. An open gas exchange System and the lkc/l2c technique were used to measure net photosynthesis and photorespiration. RuBP and ATP were determined enzymatically. The catalytic properties of RuBPCO influence net photosynthesis by determining the photorespiration/real photosynthesis ratio (PR'/RPS1). The PR'/RPS' ratio was increased as temperature during treatment of leaves in air was increased. Most of this increase was not due to the temperature dependent change of the relative solubilities of 02 and C02. The increase of the PR'/RPS' ratio was, to some extent, responsible for the relatively low temperature dependence of net photosynthe sis at air concentrations of 02 and C02. The relatively strong tempe rature dependence of the PR'/RPS' ratio of leaves from plants grown at low temperatures was related to a weaker temperature dependence of net photosynthesis. With leaves from plants grown at high temperatu res 02 inhlbition of photosynthesis showed a stronger temperature de pendence than the PR'/RPS' ratio. This indicated that net photosyn thesis was not exclusively dependent on the catalytic properties of RuBPCO but also influenced by the activation state of the enzyme. It is shown that RuBPCO in leaves pretreated at low temperatures required 20% 02 for maximum activation. Inactivation of RuBPCO due to low oxygen was related to a decrease in ATP and RuBP regeneration. However, steady state net photosynthesis was not limited by the RuBP concentration but by the activity of RuBPCO. Activation state of RuBPCO in vivo was also influenced by light in tensity, temperature and C02 concentration during treatment of leaves. The activation state increased with increasing light intensity. Where leaves were treated at low light intensity, the lower the temperature of
61 Summary The influence of the activation State of Ribulose-1.5-bisphosphate carboxylase/oxygenase (RuBPCO) and the catalytic properties of this enzyme on photosynthesis of intact leaves of white clover (Trifolium repens L.) was examined. The activation state in vivo of RuBPCO was estimated by measuring the initial carboxylase activity of rapidly prepared C02 free extracts of leaves. An open gas exchange System and the lkc/l2c technique were used to measure net photosynthesis and photorespiration. RuBP and ATP were determined enzymatically. The catalytic properties of RuBPCO influence net photosynthesis by determining the photorespiration/real photosynthesis ratio (PR'/RPS1). The PR'/RPS' ratio was increased as temperature during treatment of leaves in air was increased. Most of this increase was not due to the temperature dependent change of the relative solubilities of 02 and C02. The increase of the PR'/RPS' ratio was, to some extent, responsible for the relatively low temperature dependence of net photosynthe sis at air concentrations of 02 and C02. The relatively strong tempe rature dependence of the PR'/RPS' ratio of leaves from plants grown at low temperatures was related to a weaker temperature dependence of net photosynthesis. With leaves from plants grown at high temperatu res 02 inhlbition of photosynthesis showed a stronger temperature de pendence than the PR'/RPS' ratio. This indicated that net photosyn thesis was not exclusively dependent on the catalytic properties of RuBPCO but also influenced by the activation state of the enzyme. It is shown that RuBPCO in leaves pretreated at low temperatures required 20% 02 for maximum activation. Inactivation of RuBPCO due to low oxygen was related to a decrease in ATP and RuBP regeneration. However, steady state net photosynthesis was not limited by the RuBP concentration but by the activity of RuBPCO. Activation state of RuBPCO in vivo was also influenced by light in tensity, temperature and C02 concentration during treatment of leaves. The activation state increased with increasing light intensity. Where leaves were treated at low light intensity, the lower the temperature of
62 the leaves the higher the activation state of RuBPCO. At high light intensity temperature did not affect the activation state of RuBPCO but the light intensity which was necessary for maximum activation increa sed with temperature. With leaves from plants grown at low tempera tures RuBPCO was inactivated to a greater extent when temperature was increased during treatment at low light intensity. This suggests that maximum activation of RuBPCO in leaves from plants grown at low temperatures requires higher light intensities. Net photosynthesis and activation State of RuBPCO were closely re lated. A relatively weak temperature dependence of net photosynthesis at rate limiting light intensity was associated with a decrease in enzyme activation with increasing temperature. A weaker temperature dependence of net photosynthesis at low light intensity of leaves from plants grown at low temperatures was associated with stronger inactivation of RuBPCO with increasing temperature. A strong temperature dependence of the oxygen inhibition of net photosynthesis in leaves from plants grown at high temperatures was related to the inhibitory effect of low oxygen on RuBPCO activation at low temperatures.