In order to develop future therapeutic applications for cell penetrating peptides (CPPs), it is essential to characterize their internalization mechanisms, as they might affect the stability and the accessibility of the carried drug. Several internalization mechanisms have been described in literature, such as endocytosis and transduction. In this work we study the internalization mechanism in HeLa cells of two TIRAP derived peptides: pepTIRAP and $pepTIRAP^{ALA}$, where some of the cationic amino acids were replaced with alanines. Detailed analysis of internalization and the peptides electrostatic potential was carried out, to shed light on the internalization mechanism involved. Molecular modeling studies showed that the main difference identified between pepTIRAP and $pepTIRAP^{ALA}$ is the distribution of their electrostatic potential field. The structure of pepTIRAP displays a predominantly positive potential when compared to $pepTIRAP^{ALA}$, which has a more balanced potential distribution. In addition, docking experiments show that interactions between pepTIRAP and negatively charged molecules on the cellular surface such as heparan sulfate are stronger than the ones exhibited by $pepTIRAP^{ALA}$. A mathematical model was proposed to quantify the amount of peptide internalized or non-specifically bound to the membrane. The model indicates a stronger interaction of pepTIRAP with the plasma membrane, compared to $pepTIRAP^{ALA}$. We propose these discrepancies are related to the differences in the electrostatic potential characteristics of each peptide. In the case of pepTIRAP, these interactions lead to the formation of nucleation zones, which are the first stage of the transduction internalization mechanism. These results should be considered for effective design of a cell penetrating peptide.