Direct Interaction Between Research In Several Metal Enzymes With Metal Ions

In this paper, we studied the direct interaction between metaLloenzyme(Bacillus Subtilis Neutral Protease ,nuclease P1, alkaline phosphatase and aminoacylase I) and metal ions (Cu(II) ,Co(II)) by means of electronic paramagnetic resonance (EPR) ,visible absorption spectra and enzyme activity.All results show that additional metal ions entered the activity site replacing Zn(II) in the original active site of metalloenzyme, which influenced the enzyme activityAs for B.S.NP with only one zinc(TI) site per molecular , we found that it did produce Cu(II)- B.S.NP derivative complex and there existed two conformations of Cu(II)- B.S.NP in the active site when Cu(II) was added. These two conformations transform to each other with varied pH values.By direct interaction the added Cu(II) will enter Zn2~ Zn3 sites in the activity center of nuclease P1 which has three Zn(II) sites. However, Cu(LI) in the Zn3 site may transfer into Zn2 site with increasing pH value. In our experiment the site into which Co(II) entered at p116.0 is different with the result from reference. When Co(II) was added into enzyme solution ,the VIS spectra show the similar removal results as Cu(II).AKP from calf intestines consists of two identical subunits. After AKP interacted with Cu(II), the results indicated that Cu(II) will enter into A site of one subunit of AKP . When the pH value was raised , the Cu(II) of A site in the other subunit will transfer into B site of the subunit .Therefore , antiferromagnetism was formed by these two Cu(II) and the intensity of EPR in g1 region decreased. Added Co(I1)-AKP has the similar removal results as Cu(II).ACY from porcine kidney is a kind of dimeric protein with one Zn(II) in each subunit. The EPR of Cu(II)-ACY by direct interaction were obtained for the first time. EPR,VIS suggest that in this enzyme there also exists direct interaction.Through these works, we have expanded the investigation range of the direct interaction and found the transformation phenomena of metal ions in different active sites with varied pH values. These works will be useful for further discussion of the direct interaction between metalloenzyme and metal ions.