The Escherichia coli SlyD is a metal ion-regulated peptidyl-prolyl cis/trans-isomerase.
In Escherichia coli as many as nine different genes coding for proteins with significant homology to peptidyl-prolyl cis/trans-isomerases (PPIases) have been found. However, for three of them, the histidine-rich SlyD, the homologous gene product of ORF149, and parvulin-like SurA, it was not known whether these proteins really possess PPIase activity. To gain access to the full set of PPIases in E. coli, SlyD, the N-terminal fragment of SlyD devoid of the histidine-rich region, as well as the protein product of ORF149 of E. coli named SlpA (SlyD-like protein) were cloned, overexpressed, and purified to apparent homogeneity. On the basis of the amino acid sequences, both proteins proved to be of the FK506-binding protein type of PPIases. Only when using trypsin instead of chymotrypsin as helper enzyme in the PPIase assay, the enzymatic activity of full-length SlyD and its N-terminal fragment can be measured. For Suc-Ala-Phe-Pro-Arg-4-nitroanilide as substrate, kcat/Km of 29,600 M-1 s-1 for SlyD and 18,600 M-1 s-1 for the N-terminal fragment were obtained. Surprisingly, the PPIase activity of SlyD is reversibly regulated by binding of three Ni2+ ions to the histidine-rich, C-terminal region. Because the PPIase activity of SlpA could be established as well, we now know eight distinct PPIases with proven enzyme activity in E. coli.
Hottenrott
S
S
Forschungsstelle "Enzymologie der Proteinfaltung" der Max-Planck-Gesellschaft zur Förderung der Wissenschaften e.V., Kurt-Mothes-Str. 3, 06120 Halle/Saale, Germany.
Schumann
T
T
Plückthun
A
A
Fischer
G
G
Rahfeld
J U
JU
D000591:Amino Acid Isomerases / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009532:Nickel / Q000378:metabolism; D019696:Peptidylprolyl Isomerase; D011485:Protein Binding; D011487:Protein Conformation; D016415:Sequence Alignment; D013379:Substrate Specificity; D022021:Tacrolimus Binding Proteins
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; C085606:SlyD protein, E coli; C070683:surface layer protein A, Bacteria; C022838:nickel chloride; D009532:Nickel; D000591:Amino Acid Isomerases; D022021:Tacrolimus Binding Proteins; D019696:Peptidylprolyl Isomerase