碩士 / 國立臺灣海洋大學 / 生物科技研究所 / 100 / Spermidine/spermine acetyltransferase (Ssat), which belongs to GCN5-related N-acetyltransferase (GNAT) superfamily, catalyzes an acetyl-transferring reaction from acetyl-CoA to primary amine substrates. Two ssat isogenes, SSAT1 and SSAT2, were found in human. The substrates of human SSAT1 are spermidine and spermine, thus it regulates polyamine homeostasis. On the other hand, human SSAT2 cannot catalyze polyamine catabolism. Previous reports have indicated that thialysine can be a substrate of SSAT2. However, thialysine is rarely found in human body and its function is still unclear. Therefore, the real function of human SSAT2 is still a mystery.
Studying the evolutionary path of the properties of ssat gene family may provide some clues to elucidate the function of Ssat2. By searching the genomic database from 10 deuterostomia, we found that all vertebrates contain both ssat1 and ssat2 but invertebrates only contain ssat-like genes. Zebrafish, a lower vertebrate, contains 3 ssat1 and 2 ssat2 isogenes, which could be used as an example to trace and compare the functional evolution of Ssat2 from unicellular organism to higher animals.
We performed RT-PCR to characterize the expression pattern of ssat2a and ssat2b isogenes. ssat2a was only expressed in certain tissues but ssat2b was widely expressed in all tissues from adult fish and every stages during embryo development. When thialysine was used as a substrate, the enzyme activity was only detected from Ssat2b but not Ssat2a. Further analysis revealed that the substitution of Ser81 to Asn81 in Ssat2a was the cause of enzyme activity lost. In addition, the first 92 residues of Ssat2b were also related to the substrate binding. When 5-hydroxylysine, a structurally analog of thialysine, was used as a substrate, Ssat2b is still the only isoenzyme possessed activity. It has been reported that all members of Ssat family possess a homodimer configuration as their active form. Our results demonstrated that, in addition to homodimer, Ssat2a and Ssat2b were able to form a heterodimer with each other. Moreover, we found that Ssat2b could also form a heterodimer with all zebrafish Ssat1 isoenzymes, but Ssat2a could only form a heterodimer with zebrafish Ssat1c.
Notwithstanding the divergency of ssat2 and ssat1 from ssat ancestor gene, however, zebrafish Ssat2b preserved the activity to catalyze thialysine and its structural analogs as other Ssat-like enzyme from lower species. Therefore, such enzyme activity might be important in these species. Although Ssat2a does not show any enzyme activity, it might be able to regulate other physiological functions through protein-protein interactions.
| Identifer | oai:union.ndltd.org:TW/100NTOU5613002 |
| Date | January 2012 |
| Creators | Yu-Ling Peng, 彭于綾 |
| Contributors | Han-Jia Lin, 林翰佳 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 101 |
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