期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 99, 期 6, 页码 3469-3474出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.062043699
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资金
- Intramural NIH HHS [Z01 AG000259] Funding Source: Medline
- NCI NIH HHS [CA59717] Funding Source: Medline
- NHLBI NIH HHS [R01 HL065572, HL65572] Funding Source: Medline
- NIA NIH HHS [T32 AG000259, R37 AG009521, R01 AG009521, AG09521] Funding Source: Medline
- NICHD NIH HHS [R01 HD018179, HD18179] Funding Source: Medline
- NIGMS NIH HHS [GM08142] Funding Source: Medline
We have defined inactive alpha and omega fragments of beta-lactamase that can complement to form a functional enzyme in both bacteria and mammalian cells, serving as a readout for the interaction of proteins fused to the fragments. Critical to this advance was the identification of a tripeptide, Asn-Gly-Arg, which when juxtaposed at the carboxyl terminus of the alpha fragment increased complemented enzyme activity by up to 4 orders of magnitude. beta-Lactamase is well suited to monitoring constitutive and inducible protein interactions because it is small (29 kDa), monomeric, and assayable with a fluorescent cell-permeable substrate. The negligible background, the magnitude of induced signal caused by enzymatic amplification, and detection of signal within minutes are unparalleled in mammalian protein interaction detection systems published to date.
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