Biomedical Engineering Reference
In-Depth Information
Figure 6 shows the time-course of the synthesis of Z-Ala-Phe.OMe in 0.1M Tris-HCl
buffer pH 8.5 and ethyl acetate (50:50 ratio). As can be observed, the dipeptide concentration
decreased because of unwished by-products (Z-Ala.OMe and Z-Ala-Phe) were formed. It is
apparent that the o-methyl derivated used as nucleophile was hydrolyzed by the esterolitic
ativity of
Acacia caven
CE and then, both by-products were achieved. Further studies of
media engineering should be carried out in order to increase aminolysis/hydrolysis ratio and
other nucleophiles should also be assayed.
When Z-Gln (or Z-Phe) and Phe.OMe were used as carboxylic and nucleophilic
components, respectively, HPLC-mass spectrometry analysis of the samples indicated the
formation of dipeptide Z-Gln-Phe.OMe (or Z-Phe-Phe.OMe) after 30 min (or 1h) of reaction
(Figure 7).
2.25
ZAlaPheOMe
ZAlaPhe
ZAlaOMe
2.00
1.75
1.50
1.25
1.00
0.75
0.50
0.25
0.00
0
2
4
6
8
10
12
14
Time (h)
Figure 6. Time-course production of dipeptide Z-Ala-Phe.OMe and by-products (Z-Ala.OMe and Z-
Ala-Phe) in 50 % (v/v) ethyl acetate, using (4mM) Z-Ala as carboxylic component and (4mM)
Phe.OMe as amino nucleophilic component.
As a general conclusion, this work provides a new variety of phytoprotease as catalyst of
the peptide synthesis in aqueous-organic media. The successful synthesis of some dipeptides
has been achieved with
Acacia caven
CE although synthesis reactions need further
optimization. At present, the effect of several factors (temperature, pH, molar ratio of
enzyme/substrate, aqueous-organic media and external diffusional restrictions) on the yield of
dipeptides is being evaluated. It is interesting to emphasize that the proteolytic extract of
Acacia caven
was unable to form oligopeptides derivatives; and it showed both, a high
selectivity to form short peptide and a good specificity to react only with some amino acids.
Consequently, this enzyme constitutes a promissory catalyst for the synthesis of small
peptides.
