Structures of apo and product-bound human L-asparaginase: insights into the mechanism of autoproteolysis and substrate hydrolysis.
Structures of apo and product-bound human L-asparaginase: insights into the mechanism of autoproteolysis and substrate hydrolysis.
复制标题
apo 和产物结合的人 L-天冬酰胺酶的结构：深入了解自蛋白水解和底物水解的机制。
DOI：
10.1021/bi300870g
复制
发表时间：
2012-08-28
期刊：
影响因子：
2.9
通讯作者：
Lavie A
中科院分区：
文献类型：
--
作者：
Nomme J;Su Y;Konrad M;Lavie A
Asparaginases catalyze the hydrolysis of the amino acid asparagine to aspartate and ammonia. Bacterial asparaginases are used in cancer chemotherapy to deplete asparagine from the blood, since several hematological malignancies depend on extracellular asparagine for growth. To avoid the immune response against the bacterial enzymes it would be beneficial to replace them with human asparaginases. However, unlike the bacterial asparaginases, the human enzymes have a millimolar Km value for asparagine, making them inefficient in depleting the amino acid from blood. To facilitate the development of human variants suitable for therapeutic use, we solved the structure of human L-asparaginase (hASNase3). This asparaginase is an N-terminal nucleophile (Ntn) family member that requires autocleavage between Gly167 and Thr168 to become catalytically competent. For most Ntn-hydrolases this autoproteolytic activation occurs efficiently. In contrast, hASNas3 is relatively stable in its uncleaved state, and this allowed us to observe the structure of the enzyme prior to cleavage. To determine the structure of the cleaved state we exploited our discovery that the free amino acid glycine promotes complete cleavage of hASNase3. Both enzyme states were elucidated in the absence and presence of the product aspartate. Together, these structures provide insight into the conformational changes required for cleavage, and on the precise enzyme-substrate interactions. The new understanding of hASNase3 will serve to guide the design of variants that possess a decreased Km value for asparagine, making the human enzyme a suitable replacement for the bacterial asparaginases in cancer therapy.
登录
查看更多内容
影响因子：
--
作者：
Menniti, Miranda;Iuliano, Rodolfo;Perrotti, Nicola
通讯作者：
Perrotti, Nicola
影响因子：
1.7
作者：
Michalska, Karohna;Jaskolski, Mariusz
通讯作者：
Jaskolski, Mariusz
影响因子：
5.6
作者：
Michalska, Karolina;Bujacz, Grzegorz;Jaskolski, Mariusz
通讯作者：
Jaskolski, Mariusz
DOI：
10.1073/pnas.90.4.1474
发表时间：
1993-02-15
影响因子：
11.1
作者：
SWAIN, AL;JASKOLSKI, M;WLODAWER, A
通讯作者：
WLODAWER, A
影响因子：
11.4
作者：
Appel, I. M.;Kazemier, K. M.;Pieters, R.
通讯作者：
Pieters, R.