Molecular and Biochemical Basis for Stable Carbon Isotope Ratios in Marine Autotrophs Using Form IC and ID RubisCO

稳定碳同位素数据在生态学和地球科学中有着广泛的应用。这些关于同位素分馏的数据影响了我们对从分子到全球尺度上发生的过程的理解。过去气候的后遗症，限制当今温室气体排放命运的模型，复杂生态系统内营养转移的推断，以及酶和细胞对初级生产力控制的阐明，都使用了稳定的碳同位素分馏数据(d13C值)。Rubisco是卡尔文循环中的二氧化碳固定酶，是迄今为止地球上最重要的催化固碳的酶，被光自养和化学自养广泛使用。目前，在自养生物中已经描述了五种形式的Rubisco：形式IA-D和形式II，它们的结构和催化性能不同。至少有4种形式对海洋初级生产力有显著贡献。到目前为止，只测定了一种形式的IA Rubisco(文蛤Solemya的化学自养共生菌；24.5.)、两种形式的IB酶(菠菜和聚球藻PCC6301；分别为29和22)和两种形式的第二种酶(红色红螺菌和厚壁Riftia pachyptila共生菌；~18.对于两者)RuBisCOs的动力学同位素效应(Kies)，即C固定的d13C与所使用的CO2底物的不同程度。确定不同RuBisCO的Kie对于理解、解释和模拟海洋学和全球碳的d13C值至关重要。PI正在评估IC型和ID RuBisCOs的Kie，以估计这些酶的全部同位素分馏范围。Rubisco酶是从具有代表性的海洋浮游植物和自养细菌中分离和纯化的。他们正在研究来自模式物种球形红杆菌和海洋分离株S185-9A1的IC Rubisco，以及来自浮游球虫Emily huxleyi和硅藻中肋骨条藻的ID Rubisco。分离的蛋白质正在根据氨基酸序列、组成、Vmax和Km(CO2)进行特征分析。最后，用高精度的方法测定了动力学同位素效应。

Stable carbon isotope data are used widely in ecology and earth sciences. These data on isotope fractionations influence our understandings of processes occurring on molecular to global scales. Hindcasts of past climates, models constraining the fates of present day greenhouse gas emissions, inferences of trophic transfers within complex ecosystems, and elucidation of enzyme and cellular controls on primary productivity have all used stable carbon isotope fractionation data (d13C values). RubisCO, the CO2-fixing enzyme for the Calvin Cycle, is by far the most important enzyme catalyzing carbon-fixation on earth, used widely by photoautotrophs and chemoautotrophs. Presently, five forms of RubisCO have been described in autotrophs: Forms IA-D, and Form II, which differ in their structure and catalytic properties. At least 4 forms contribute significantly to primary productivity in the ocean. To date, the kinetic isotope effects (KIEs) for RubisCOs, the degree to which the d13C of the C fixed differs from the CO2 substrate used, have only been determined for one form IA RubisCO (chemoautotrophic symbionts of the clam Solemya; 24.5.), two form IB enzymes (spinach and Synechococcus PCC 6301; 29 and 22., respectively) and two form II's (Rhodospirillum rubrum and Riftia pachyptila symbionts; ~18. for both). Determining the KIEs of different RubisCOs is critical to understanding, interpreting, and modeling of d13C values in oceanography and global carbon. The PIs are assessing the KIEs for form IC and ID RubisCOs, to estimate the full range of isotope fractionations by these enzymes. The RubisCO enzymes are being isolated and purified from representative marine phytoplankton and autotrophic bacteria. They are studying form IC RubisCO from the model species Rhodobacter sphaeroides and marine isolate S185-9A1, and form ID RubisCO from the coccolithophore Emiliania huxleyi and the diatom Skeletonema costatum. The isolated proteins are being characterized with respect to amino acid sequence, composition, Vmax, and Km (CO2). Finally, the kinetic isotope effects are being determined by high precision methods.