A single-step process to polymer nanofiber meshes that possess biofunctional peptide segments on their surfaces is described here, which requires a standard electrospinning setup only. Spinning a homogeneous mixture composed of a valuable polymer-peptide conjugate (poly(lactic acid)-block-CGGRGDS) and a biocompatible commodity poly(lactic-co-glycolic acid) (PLGA) leads to nonwovens where the bioactive peptide part is enriched up to 11 times on their fiber surface. This is determined by X-ray photoelectron spectroscopy (XPS). The surface accessibility of the peptide is proved on the macroscale by contact angle measurements comparing pure PLGA fibers with GRGDS-functionalized fiber meshes as well as on the nanoscale by probing electrostatic interaction between CGGRGDS surface functionalities and a colloidal silica probe via atomic force microscopy (AFM). Ultimately, bioavailability and bioactivity of the peptides on the fiber surfaces are demonstrated, showing that the meshes promote adhesion and migration of fibroblasts in comparison to pure PLGA meshes. The one-step production of hydrophilic PLGA-based fibers could be exploited to electrospin into living cell culture without indication of toxic adverse effects on cell proliferation. This might be useful for directly production of cell-loaded scaffolds or biohybrid materials.