Fumarase (Fumarate hytdratase) is an enzyme that catalyzes the reversible hydration/dehydration of fumarate to S-malate and is involved in the tricarboxylic acid (TCA), or Krebs cycle. This enzyme exists in both a cytosolic form and an N-terminal extended mitochondrial form. The N-terminal extended form is targeted to the mitochondrion, where the removal of the extension is the same form as in the cytoplasm. Fumarase deficiency can lead to progressive encephalopathy, cerebral atrophy and developmental delay and this enzyme also is thought to act as a tumor suppressor. Recombinant protein corresponding to aa44-510 from human Fumarase, expressed in E. coli. Accession: NP_000134 Molecular Weight: ~50.2kD (467aa) Specific Activity: >25unit/mg, and is defined as the amount of enzyme that cleaves 1umole of L-Malate to Fumarate/minute at pH 7.5 at 37C. Activity Assay: 1. Prepare a 180ul reaction mix into a suitable container: The concentrations are 90mM potassium phosphate, 90 mM L-Malic acid. 2. Equilibrate to 37C and monitor at A260nm until the value is constant using a spectrophotometer. 3. Add 20ul of recombinant Fumarase protein 50ug/ml in assay buffer. 4. Mix by inversion and record the increase at A260nm for 5 minutes. Amino Acid Sequence: MASQNSFRIE YDTFGELKVP NDKYYGAQTV RSTMNFKIGG VTERMPTPVI KAFGILKRAA AEVNQDYGLD PKIANAIMKA ADEVAEGKLN DHFPLVVWQT GSGTQTNMNV NEVISNRAIE MLGGELGSKI PVHPNDHVNK SQSSNDTFPT AMHIAAAIEV HEVLLPGLQK LHDALDAKSK EFAQIIKIGR THTQDAVPLT LGQEFSGYVQ QVKYAMTRIK AAMPRIYELA AGGTAVGTGL NTRIGFAEKV AAKVAALTGL PFVTAPNKFE ALAAHDALVE LSGAMNTTAC SLMKIANDIR FLGSGPRSGL GELILPENEP GSSIMPGKVN PTQCEAMTMV AAQVMGNHVA VTVGGSNGHF ELNVFKPMMI KNVLHSARLL GDASVSFTEN CVVGIQANTE RINKLMNESL MLVTALNPHI GYDKAAKIAK TAHKNGSTLK ETAIELGYLT AEQFDEWVKP KDMLGPK Storage and Stability: May be stored at 4C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20C. Aliquots are stable for 6 months after receipt at -20C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
