Sequence Analysis and Chromosomal Localization of Human Cap Z: CONSERVED RESIDUES WITHIN THE ACTIN-BINDING DOMAIN MAY LINK CAP Z TO GELSOLIN/SEVERIN AND PROFILIN PROTEIN FAMILIES

Emily A Barron-Casella,Michelle A Torres,Stephen W Scherer,Henry H.Q Heng,Lap-Chee Tsui,James F Casella

doi:10.1074/jbc.270.37.21472

Abstract

From a human retinal cDNA library, we have isolated cDNAs that are homologs for the alpha 2 and beta subunits of chicken Cap Z. The derived human alpha subunit shares 95% amino acid identity with the chicken alpha 2 subunit; the beta subunit is 99% identical to the chicken subunit residues 1-243. The remaining portion of the human beta subunit (244-272) diverges significantly with only 8 out of 29 C-terminal amino acids conserved between the two species. This lack of conservation is of particular interest because the chicken C terminus contains an actin-binding domain. Cosedimentation assays with F-actin show that human Cap Z binds actin with an affinity equal that of chicken Cap Z. These results point to the eight shared amino acids as critical for actin binding, three of which are regularly spaced leucines. These apolar residues and one outside the region of divergence align well with those residues of the actin-binding alpha-helix proposed for gelsolin segment 1. The apolar residues as well as three polar amino acids are also conserved in other capping, capping and severing, and monomer-binding proteins. Amino acid substitutions in the chicken beta subunit of the two most highly conserved leucines result in significant decreases in F-actin binding activity. The human alpha 2 gene (CAPZA2) has been mapped to chromosome 7 position q31.2-q31.3 and the beta gene (CAPZB) to chromosome 1 region p36.1.

Highlights

From a human retinal cDNA library, we have isolated cDNAs that are homologs for the ␣2 and ␤ subunits of chicken Cap Z
Nucleotide and Predicted Amino Acid Sequence—Several human cDNAs were isolated from a human retinal library for both the Cap Z ␣ and ␤ subunits
Sequence Comparison between Human and Chicken Cap Z—Our results show that the majority of human Cap Z is over 95% identical to the ␣2␤ heterodimer of chicken Cap Z

Summary

Introduction

From a human retinal cDNA library, we have isolated cDNAs that are homologs for the ␣2 and ␤ subunits of chicken Cap Z. The remaining portion of the human ␤ subunit (244 –272) diverges significantly with only 8 out of 29 C-terminal amino acids conserved between the two species This lack of conservation is of particular interest because the chicken C terminus contains an actin-binding domain. Cosedimentation assays with F-actin show that human Cap Z binds actin with an affinity equal that of chicken Cap Z These results point to the eight shared amino acids as critical for actin binding, three of which are regularly spaced leucines. To confirm the functional importance of these conserved residues, we have made amino acid substitutions for two of the most conserved residues in the ␤ domain of chicken Cap Z and studied the F-actin binding characteristics of the heterodimers (␣1/mutated ␤) in actin cosedimentation and binding assays

Methods

Results

Conclusion