Molecular cloning and expression in Escherichia coli of a cDNA encoding human pancreatic elastase 2.
We have cloned a DNA that is complementary to the messenger RNA that encodes human pancreatic elastase 2 from a human pancreatic cDNA library using a cloned cDNA for rat pancreatic elastase 2 messenger RNA. This complementary DNA contains the entire protein coding region of 807 nucleotides which encodes preproelastase of 269 amino acids, and 4 and 82 nucleotides of the 5'- and 3'-untranslated sequences, respectively. When this deduced amino acid sequence was compared with known amino acid sequences it showed 82% homology with rat pancreatic elastase 2. This deduced sequence also contains a 16-amino-acid peptide identical with the N-terminal sequence determined for native human pancreatic proelastase 2. Taking the above findings together, we conclude that the cloned cDNA encodes a mature enzyme of 241 amino acids including 16 and 12 amino acids for a signal peptide and an activation peptide, respectively. Moreover, the predicted key amino acid residues involved in determining the substrate specificity of mammalian pancreatic elastase 2 are retained in the human enzyme. Cloned human pancreatic elastase 2 cDNA was expressed in E. coli as a mature and pro-form protein. Both resulting proteins showed immunoreactivity toward anti-elastase serum and enzymatic activity. We have also cloned and sequenced a porcine pancreatic elastase 2 cDNA.
Shirasu
Y
Y
Kirin Brewery Co., Ltd., Pharmaceutical Laboratory, Gunma.
Yoshida
H
H
Matsuki
S
S
Takemura
K
K
Ikeda
N
N
Shimada
Y
Y
Ozawa
T
T
Mikayama
T
T
Iijima
H
H
Ishida
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D010179:Pancreas / Q000201:enzymology*; D010196:Pancreatic Elastase / Q000096:biosynthesis / Q000235:genetics*; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D012689:Sequence Homology, Nucleic Acid; D013552:Swine; D014170:Transformation, Genetic
D004274:DNA, Recombinant; D012333:RNA, Messenger; D004247:DNA; D004262:DNA Restriction Enzymes; D010196:Pancreatic Elastase