Discerning how different major groups of organisms are related to each other and tracing their evolution from the common ancestor remains controversial and unsolved. In recent years, much new information based on a large number of gene and protein sequences has become available. So far phylogenetic analysis can be carried out based on either nucleic acid or protein sequences. However it was become evident that both approaches have a many serious limitations and pitfalls. Our novel findings follows from analysis of loops in elongation factors EF1A using novel informative characteristic which was called “loops” method. The method is based on the ability of amino acid sequences form loops in protein structure. The specificity of a criterion for grouping organisms is distinctly evident from the analysis of the loops using EF1A for three kingdoms of life. Each kingdom displayed variations in the number of loops and their location within the three EF1À domains and can be consider as imprint of molecular evolution. Stronger support that animals and fungi are sibling kingdoms was found.