Recombinant Human NF-L Rod 80-400

Human NF-L sequence was based on that was NP_006149.2 which was inserted into the eukaryotic expression vector pET30a(+) which adds an N terminal His-tag and some other sequence, underlined below. This sequence includes a thrombin cleavage site (blue), an S-tag affinity peptide (red) and an enterokinase cleavage site (green). This construct is designed to express the entire α-helical “rod” region without the globular “head” or “tail” regions.

MHHHHHHSSG LVPRGSGMKE TAAAKFERQH MDSPDLGTDD DDKAMADIGS EFTQEKAQLQ  60
DLNDRFASFI ERVHELEQQN KVLEAELLVL RQKHSEPSRF RALYEQEIRD LRLAAEDATN 120
EKQALQGERE GLEETLRNLQ ARYEEEVLSR EDAEGRLMEA RKGADEAALA RAELEKRIDS 180
LMDEISFLKK VHEEEIAELQ AQIQYAQISV EMDVTKPDLS AALKDIRAQY EKLAAKNMQN 240
AEEWFKSRFT VLTESAAKNT DAVRAAKDEV SESRRLLKAK TLEIEACRGM NEALEKQLQE 300
LEDKQNADIS AMQDTINKLE NELRTTKSEM ARYLKEYQDL LNVKMALDIE IAAYRKLLEG 360
EETRL 365

Number of amino acids: 365
Molecular weight: 41898.93
Theoretical pI: 4.97

Amino acid composition:
Ala (A) 44 12.1%
Arg (R) 27 7.4%
Asn (N) 12 3.3%
Asp (D) 25 6.8%
Cys (C) 1 0.3%
Gln (Q) 22 6.0%
Glu (E) 53 14.5%
Gly (G) 11 3.0%
His (H) 10 2.7%
Ile (I) 14 3.8%
Leu (L) 42 11.5%
Lys (K) 27 7.4%
Met (M) 12 3.3%
Phe (F) 8 2.2%
Pro (P) 4 1.1%
Ser (S) 19 5.2%
Thr (T) 14 3.8%
Trp (W) 1 0.3%
Tyr (Y) 7 1.9%
Val (V) 12 3.3%

Total number of negatively charged residues (Asp + Glu): 78
Total number of positively charged residues (Arg + Lys): 54

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 15930
Abs 0.1% (=1 g/l) 0.380, assuming all pairs of Cys residues form cystines

Ext. coefficient 15930
Abs 0.1% (=1 g/l) 0.380, assuming all Cys residues are reduced