A cold-regulated protein from bermudagrass crowns is a chitinase

Sodium dodecylsulfate (SDS) polyacrylamide gel electrophoresis (PAGE) and fluorography of in vivo radiolabeled proteins has revealed that protein synthesis in crowns of 'Midiron' and 'Tifgreen' bermudagrasses (Cynodon dactylon L. Pers. x C. transvaalensis Burtt-Davy) is altered in association with cold acclimation (CA). Increased synthesis of some proteins in association with CA in Midiron crowns correlates with the greater freezing tolerance of Midiron compared with Tifgreen. This study was undertaken to (i) characterize further alterations in protein synthesis in Midiron and Tifgreen crowns utilizing analytical isoelectric focusing (IEF)/SDS-PAGE [two-dimensional (2D-) PAGE], and (ii) identify cold-regulated (COR) proteins, proteins encoded by genes persistently upregulated during CA, through sequence analysis of peptides recovered after micropreparative 2D-PAGE and protease digestion. Plants were grown for 26 d in controlled-environment chambers under acclimating [8/2 degrees C (day/night) cycles with 10 h photoperiods] or nonacclimating [28/24 degrees C] conditions. Proteins synthesized by isolated crowns were radiolabeled in vivo for 16 h with S-35-methionine and S-35-cysteine. Among several groups of COR proteins synthesized in Midiron and Tifgreen crowns, low molecular weight (about 20-28 kilodalton) basic (LB) COR proteins were synthesized in greater amounts and numbers in Midiron than Tifgreen crowns. Among the LB COR proteins, a 27-kD basic COR protein was synthesized prominently in crowns of both Midiron and Tifgreen plants. This protein from Midiron crowns was designated COR27 and identified as a chitinase (E.C. 3.2.1.14) through protein sequencing.