A highly sensitive photometric method for the quantitation of protein separated in SDS-PAGE micro-slab gels was used to determine the relative amounts of rubisco in palisade, spongy and guard cells of Vicia faba L. Cell types were obtained by dissection from freeze-dried leaf or by protoplast isolation. Protein profiles of palisade and spongy mesophyll were virtually identical. The rubisco large subunit (ls) was the most abundant protein in both mesophyll cell types (32-35%, protein basis). A comparison of mesophyll and guard-cell protein profiles indicated considerable differences. For guard cells from lyophilized tissue, the maximum possible contribution of the rubisco ls to total protein was 1.08%. Among guard-cell protoplast preparation, the maximum contribution of the rubisco ls was variable and greater (2-5%, protein basis), which is consistent with contamination by mesophyll. On a cell basis, these data indicate that the upper limit for guard-cell rubisco is 0.2% of that of mesophyll. These findings indicate that rubisco activity in guard cells is negligible, if it is present at all. / Kinetic properties of phosphoenolpyruvate carboxylase (PEPC) of guard cells of Vicia faba L. in different stage of stomatal opening were determined at two pHs and in the presence of malate. PEPC was "extracted" from individually excised cells in microdroplets of reagent. After fewer than 15 second, the reaction progress was monitored in real-time. Dephosphorylation of phosphoenolpyruvate (PEP) by endogenous phosphatase activity was corrected for by parallel assays with no Mg$\sp{2+}$. A pH increase from 7.0 to 8.5 decreased Km(PEPMg) 3.5-fold. Malate inhibited Km(PEPMg) at both pHs, but inhibition was 3-fold greater at pH 7.0. These data indicate that the combined effects of pH and malate can produce changes in enzyme velocity by more than 10-fold. However, PEPC Km(PEPMg) and maximum enzyme velocities of guard cells of closed, opening, and open stomata were affected similarly by pH and malate. These findings are consistent with a role of PEPC in cytoplasmic pH maintenance in guard cells. We discuss the physiological implications of the kinetics of PEPC in relation to guard-cell function. / Source: Dissertation Abstracts International, Volume: 50-03, Section: B, page: 0817. / Major Professor: William H. Outlaw. / Thesis (Ph.D.)--The Florida State University, 1989.
| Identifer | oai:union.ndltd.org:fsu.edu/oai:fsu.digital.flvc.org:fsu_77992 |
| Contributors | Tarczynski, Mitchell C., Florida State University |
| Source Sets | Florida State University |
| Language | English |
| Detected Language | English |
| Type | Text |
| Format | 112 p. |
| Rights | On campus use only. |
| Relation | Dissertation Abstracts International |
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