Nanohybrids based on biomolecular nanostructures and graphene quantum dots (GQDs) have found wide application in the biological and biomedical fields. Herein, the design of a peptide with trifunctional motifs is reported as the precursor building block for constructing a novel multifunctional protein nanofiber (PNF), and further conjugated with highly fluorescent GQDs by noncovalent interactions. The physicochemical properties of these PNF-GQD nanohybrids are thoroughly characterized by a variety of spectroscopic and microscopic techniques, revealing that the GQDs essentially maintain their favorable optical properties in the nanohybrids. A good biocompatibility of the PNF-GQD nanohybrids is found with cell viability assays. With both, a recognition moiety (RGD) and an imaging probe (GQD), these PNF-GQD nanohybrids possess the capability of targeting and imaging tumor cells simultaneously. A potential application of these novel nanohybrids, i.e., fluorescence imaging of HeLa tumor cells, has been investigated by confocal fluorescence microscopy, which shows much enhanced labeling efficiency compared with GQDs only. Moreover, cellular internalization by nontumorous COS-7 cells was much weaker than by HeLa cells. Our results show that GQD-decorated PNF nanohybrids have great potential as multifunctional platforms for biomedical applications, particularly, where the capability of sensitive tracking and efficient labeling is appreciated.