Degradation of myoglobin by polymeric artificial metalloproteases containing catalytic modules with various catalytic group densities: site selectivity in peptide bond cleavage.
Mononuclear, dinuclear, and tetranuclear artificial metalloproteases were prepared by attaching respective catalytic modules containing the Cu(II) complex of cyclen (Cu(II)Cyc) to a derivative of cross-linked polystyrene. The polymeric artificial metalloproteases effectively cleaved peptide bonds of myoglobin (Mb) by hydrolysis. The proteolytic activity increased considerably as the catalytic group density was raised: the ratio of k(cat)/K(m) was 1:13:100 for the mono-, di-, and tetranuclear catalysts. In the degradation of Mb by the dinuclear catalyst, two pairs of intermediate proteins accumulated. One of the two initial cleavage sites leading to the formation of the protein fragments is identified as Gln(91)-Ser(92) and the other is suggested as Ala(94)-Thr(95). On the basis of a molecular modeling study by using the X-ray crystallographic structure of Mb, the site-selectivity is attributed to anchorage of one Cu(II)Cyc unit of the catalytic module to a heme carboxylate of Mb. The high site selectivity for the initial cleavage of a protein substrate and mechanistic analysis of the catalytic action are unprecedented for polymeric artificial enzymes.
Yoo
Chang Eun
CE
Department of Chemistry, Seoul National University, Seoul 151-747, Korea.
Chae
Pil Seok
PS
Kim
Jung Eun
JE
Jeong
Eui June
EJ
Suh
Junghun
J
D000818:Animals; D040761:Biomimetic Materials / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D003300:Copper / Q000737:chemistry; D006736:Horses; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D045726:Metalloproteases / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D009211:Myoglobin / Q000737:chemistry* / Q000378:metabolism; D009942:Organometallic Compounds / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013329:Structure-Activity Relationship
D009211:Myoglobin; D009942:Organometallic Compounds; D003300:Copper; D045726:Metalloproteases