Substrate Specificity of the Sialic Acid Biosynthetic Pathway
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- 作者：Christina L. Jacobs ; Scarlett Goon ; Kevin J. Yarema ; Stephan Hinderlich ; Howard C. Hang ; Diana H. Chai ; and Carolyn R. Bertozzi
- 刊名：Biochemistry
- 出版年：2001
- 出版时间：October 30, 2001
- 年：2001
- 卷：40
- 期：43
- 页码：12864 - 12874
- 全文大小：158K
- 年卷期：v.40,no.43(October 30, 2001)
- ISSN：1520-4995
文摘
Unnatural analogues of sialic acid can be delivered to mammalian cell surfaces through themetabolic transformation of unnatural N-acetylmannosamine (ManNAc) derivatives. In previous studies,mannosamine analogues bearing simple N-acyl groups up to five carbon atoms in length were recognizedas substrates by the biosynthetic machinery and transformed into cell surface sialoglycoconjugates [Keppler,O. T., et al. (2001) Glycobiology 11, 11R-18R]. Such structural alterations to cell surface glycans can beused to probe carbohydrate-dependent phenomena. This report describes our investigation into the extentof tolerance of the pathway toward additional structural alterations of the N-acyl substituent of ManNAc.A panel of analogues with ketone-containing N-acyl groups that varied in the length or steric bulk waschemically synthesized and tested for metabolic conversion to cell surface glycans. We found that extensionof the N-acyl chain to six, seven, or eight carbon atoms dramatically reduced utilization by the biosyntheticmachinery. Likewise, branching from the linear chain reduced metabolic conversion. Quantitation ofmetabolic intermediates suggested that cellular metabolism is limited by the phosphorylation of theN-acylmannosamines by ManNAc 6-kinase in the first step of the pathway. This was confirmed byenzymatic assay of the partially purified enzyme with unnatural substrates. Identification of ManNAc6-kinase as a bottleneck for unnatural sialic acid biosynthesis provides a target for expanding the metabolicpromiscuity of mammalian cells.