Purification, molecular and enzymic characterization of an acid RNase from the insect Ceratitis capitata

Authors


  • Definition. Fick, unit of diffusion equivalent to 10−7 cm2· s−1.

  • Note. Portions of this paper (including Materials and Methods, Tables 1 and 3, and Figs 1, 5, 6 and 7) are presented in miniprint at the end of the paper.

Correspondence to J. M. García-Segura, Departamento de Bioquimica, Facultad de Ciencias Quimicas, Universidad Complutense de Madrid, E-28040 Madrid, Spain

Abstract

An acid ribonuclease has been purified from the insect Ceratitis capitata. The specific activity of the purified enzyme is 580 units/mg. This enzyme is a single polypeptide chain of about 35.5 kDa, containing only one disulfide bridge and no free – SH groups. The A0.1%1 cm at 280 nm is 1.90. The hydrodynamic radius of the native enzyme is 2.5 nm. The secondary structure of this RNase is composed of 10%α-helix, 31%β-structure and 59% aperiodic conformation with an average number of residues per helical segment of 10, based on circular dichroic measurements. Optimum parameters for the enzyme activity are pH 5.5, 0.15 M ionic strength and 40°C. Divalent cations are not required for the enzymic catalysis. This enzyme has been characterized as cyclizing endoribonuclease.

Abbreviations
RNase

ribonuclease

SDS

sodium dodecyl sulphate

pI

isoelectric point

poly(A)

poly(adenylic acid)

poly(C)

poly(cytidyclic acid)

poly(G)

poly(guanylic acid)

poly(U)

poly(uridylic acid)

Enzyme
 

Acid ribonuclease from C. captita (EC 3.1.27.–)

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